Received: 7 November 2018 Revised: 21 January 2019 Accepted: 21 January 2019

DOI: 10.1002/yea.3380

ECOYEAST REVIEW

Bioprospecting for brewers: Exploiting natural diversity for

naturally diverse beers

Francisco A. Cubillos1,2 |Brian Gibson3 | Nubia Grijalva‐Vallejos4 |

Kristoffer Krogerus3,5 | Jarkko Nikulin3,6

1
Departamento de Biología, Facultad de
Química y Biología, Universidad de Santiago Abstract
de Chile, Santiago, Chile The burgeoning interest in archaic, traditional, and novel beer styles has coincided
2
Millennium Institute for Integrative Biology
with a growing appreciation of the role of yeasts in determining beer character as well
(iBio), Santiago, Chile
3
Industrial Biotechnology and Food Solutions,
as a better understanding of the ecology and biogeography of yeasts. Multiple studies
VTT Technical Research Centre of Finland Ltd, in recent years have highlighted the potential of wild Saccharomyces and non‐Saccha-
Espoo, Finland
4
romyces yeasts for production of beers with novel flavour profiles and other desirable
Institute for Integrative Systems Biology
(I2SysBio), University of Valencia‐CSIC, properties. Yeasts isolated from spontaneously fermented beers as well as from other
Valencia, Spain food systems (wine, bread, and kombucha) have shown promise for brewing applica-
5
Department of Biotechnology and Chemical
tion, and there is evidence that such cross‐system transfers have occurred naturally in
Technology, Aalto University, School of
Chemical Technology, Espoo, Finland the past. We review here the available literature pertaining to the use of nonconven-
6
Chemical Process Engineering, Faculty of tional yeasts in brewing, with a focus on the origins of these yeasts, including
Technology, University of Oulu, Oulu, Finland
methods of isolation. Practical aspects of utilizing nondomesticated yeasts are
Correspondence discussed, and modern methods to facilitate discovery of yeasts with brewing poten-
Brian Gibson, VTT Technical Research Centre
of Finland Ltd, Tietotie 2, P.O. Box 1000, FI‐ tial are highlighted.
02044 VTT, Espoo, Finland.
Email: brian.gibson@vtt.fi KEY W ORDS

Funding information beer, domestication, flavour, identification, isolation, yeast

Alfred Kordelinin Säätiö; Suomen
Kulttuurirahasto; Svenska Kulturfonden;
Fonds Baillet Latour; Academy of Finland,
Grant/Award Numbers: 305453 and 276480;
Secretaría de Educación Superior, Ciencia,
Tecnología e Innovación; Instituto Milenio
iBio ‐ Iniciativa Científica Milenio MINECON

1 | I N T RO D U CT I O N
Brewing yeasts have a direct influence on the character and quality of
beer, with specific types being used for the production of particular
styles. Different yeasts, typically strains of Saccharomyces cerevisiae,
are used for production of Kölsch, maltøl, sahti, saison, Weissbier, and
so forth (White & Zainasheff, 2010). Although the properties of the dif-
ferent groups vary, they normally share characteristic domestication
All authors made an equal contribution to this manuscript.
traits such as strong flocculation, efficient sugar use, and lack of off‐
flavour production (Gallone et al., 2016; Gonçalves et al., 2016).
Though limited to specific beer styles, yeasts belonging to other genera,
such as Brettanomyces bruxellensis, can also make significant contribu-
tions to beer character and also exhibit signs of adaptation to the
brewing environment (Crauwels et al., 2015). The contribution of yeast
strains to product quality is often overlooked, but, arguably, strain has
as much influence on beer character as raw materials.
Prior to industrialization of brewing, it may be speculated that
there existed greater diversity within the brewing yeast group, with

Yeast. 2019;36:383–398. wileyonlinelibrary.com/journal/yea © 2019 John Wiley & Sons, Ltd. 383
384 CUBILLOS ET AL.

individual strains being maintained by individual breweries and even

households. Centralized beer production may have had the effect of Outstanding questions related to bioprospecting for
reducing the number of strains used for brewing, particularly those brewing
strains used for lager beer production. Microbiological developments
Saccharomyces eubayanus has not yet been isolated in
in the 19th century and, in particular, the use of pure yeast cultures
Europe despite the presumed European provenance of
pioneered by Christian Hansen in the 1880s greatly improved the con-
Saccharomyces pastorianus (Saccharomyces
sistency and quality of beer but would have further limited the num-
cerevisiae × S. eubayanus). The species, though probably
ber of strains in common use. Hansen also adopted a policy of
present (Alsammar et al., 2018), is clearly not abundant in
donating the Carlsberg Brewery's lager yeast strains to other brewing
Europe, and the circumstances leading to the lager yeast
companies for their unrestricted use (Barnett & Lichtenthaler, 2001). It
hybridization remain unclear. It remains to be seen if
is probable that much phenotypic diversity was lost during this period.
European S. eubayanus occupies a very specific niche or
Norwegian farmhouse “kveik” ale yeasts, which presumably survived
has a more widespread, but sparse, distribution, either of
this period unscathed, show several points of phenotypic divergence
which might explain its scarcity. Another scenario involving
compared to commercial brewing yeast strains and appear to be a
a non‐European hybridization, possibly in Eastern Asia,
hybrid lineage formed from a union between one of the main ale
where the species is more commonly found, followed by
brewing yeast lineages and an unknown, possibly wild yeast lineage
transport to Europe, could be explored.
(Preiss et al., 2018). It is possible that there still remain similar pockets
Although the brewing potential of S. eubayanus has been
of diversity around the world in traditional brewing systems that were
clearly established, there are limited, or no, reports yet of
not influenced by the presumed industrial bottleneck.
the potential application of the other maltose‐positive
The continued success in the craft beer market (Garavaglia &
species within the genus, for example, Saccharomyces
Swinnen, 2018) and consumer interest in varied beer styles (Aquilani,
paradoxus, Saccharomyces uvarum, and Saccharomyces jurei.
Laureti, Poponi, & Secondi, 2015) has stimulated interest in the applica-
The kveik yeasts used for traditional Norwegian farmhouse
tion of new yeasts in brewing (for reviews, see Gibson et al., 2017;
brewing have recently been recognized as a genetically and
Hittinger, Steele, & Ryder, 2018). Such yeasts include the wild yeast Sac-
phenotypically distinct group of domesticated brewing
charomyces eubayanus, a recently discovered parent of the lager yeast
yeasts (Preiss, Tyrawa, Krogerus, Garshol, & Van Der
hybrid, as well as numerous and diverse species including Torulaspora
Merwe, 2018). Whether other “lost” brewing yeast lineages
delbrueckii, Lachancea thermotolerans, and Mrakia gelida (De Francesco
exist associated with traditional brewing practices is not
et al., 2018; Domizio, House, Joseph, Bisson, & Bamforth, 2016; Michel
known.
et al., 2016). Compared with conventional brewing yeast, such species
Yeasts with potential application in brewing have been
offer a number of functional benefits, including novel flavour profiles,
isolated from cacao, kombucha, and sourdough cultures
production of flavourful low‐alcohol beers, reduction of calorie content,
(Bellut et al., 2018; Holt, Mukherjee, Lievens, Verstrepen,
and primary souring, and can be used to enhance existing beer styles, cre-
& Thevelein, 2018; Mascia et al., 2015). Yeast strains
ate new styles, or faithfully recreate traditional or ancient beer styles. In
isolated from many other food fermentation systems could
other words, the use of such yeasts has the potential to differentiate
be likewise be exploited for production of speciality beers.
products in an already diverse market.
Artificial domestication has been applied to S. eubayanus
A wide range of strategies is nowadays available for providing
strains to enhance their industrial applicability. Similar
new yeasts for the fermentation industry. These include experimental
approaches could be explored with members of the genus
evolution, mutagenesis, breeding, and yeast isolation from the wild
as well as non‐Saccharomyces yeasts with traits of interest
(Steensels, Meersman, Snoek, Saels, & Verstrepen, 2014; Steensels,
for brewing.
Snoek, et al., 2014). The latter option represents one of the most
attractive alternatives for brewers: (a) It takes advantage of the natural
biodiversity of the microorganisms and the habitat properties found
within a geographical region, (b) new species and strains can incorpo- other food and beverage systems. We also discuss the potential of in

rate novel flavours and aromas into the fermentation process, (c) some silico approaches for identification of yeasts in the “postgenome” era.

yeasts isolated from the wild already have Generally Recognized as

Safe/Qualified Presumption of Safety status, and (d) current regula-
tions favour the utilization of nonmodified genetic stocks. In this way,
identity and uniqueness are added to differentiate the production line.
Alternative yeasts with brewing potential have been isolated from
various environmental niches. Here, we review the use of atypical
yeasts in brewing, the varied origins of these yeasts, and the modern
techniques used for isolation and identification. Also, considered are
the few examples where brewing yeast has been commandeered from
2 | ISOLATION AND BREWING POTENTIAL
OF WILD S A C C H A R O M Y C E S S P P .
2.1 | Methods for isolating yeasts from the wild
Recent efforts to expand the repertoire of yeasts available for brewing
have focused on the isolation of new Saccharomyces strains or species
CUBILLOS ET AL. 385

from the wild across the globe (Bing, Han, Liu, Wang, & Bai, 2014;
Gayevskiy & Goddard, 2016; Libkind et al., 2011; Naseeb et al.,
2018; Peris et al., 2014; Figure 1). For this, a series of methods have
been proposed with different yields in terms of the genus and species
obtained depending on the selection method utilized (Sampaio &
Gonçalves, 2008; Sniegowski, Dombrowski, & Fingerman, 2002).
Saccharomyces yeasts are considered ethanol resistant and are suitable
for an extensive set of fermentation processes, including wine, beer,
sake, and bioethanol (Cubillos, 2016; Legras, Merdinoglu, Cornuet, &
Karst, 2007; Steensels, Snoek, et al., 2014), all of them containing high
ethanol levels (above 4% v/v). One of the most cited methods for suc-
cessfully isolating Saccharomyces yeasts from the wild corresponds to

FIGURE 1 Sampling from Nothofagus trees

in Chile. (a) Nothofagus leaves found on snow
from Nothofagus dombeyi (left) and Nothofagus
pumilio (right) and (b) isolation procedure. In
this picture, Francisco Cubillos and Gilles
Fischer, (c) incision performed in the N. pumilio
tree to obtain bark samples and (d) collection
tubes containing yeast nitrogen base,
raffinose, and ethanol 8% [Colour figure can
be viewed at wileyonlinelibrary.com]
386
media containing yeast extracts and sucrose supplemented with 7.6%
(v/v) of ethanol and that was originally proposed by Sniegowski et al.
(2002). This recipe allowed the isolation of dozens of S. cerevisiae and
Saccharomyces paradoxus strains from soil samples around oaks
(Quercus spp.), demonstrating that S. cerevisiae isolates can also be
obtained from nonfermentative and non‐industrial settings
(Sniegowski et al., 2002). In this way, for the past 10 years, a series
of new Saccharomyces species have been recovered from bark,
decayed leaves, and soil samples utilizing this recipe. These include
Saccharomyces arboricola (S. A. Wang & Bai, 2008) and Saccharomyces
jurei (Naseeb et al., 2018). Because Saccharomyces species have a clear
preference for hexoses, such as glucose, galactose, and fructose, or
simple oligosaccharides, such as maltose, sucrose, and raffinose, most
isolation media contain at least one of these sugars. Thus, modifica-
tions of the original recipe included yeast nitrogen base instead of
yeast extract and raffinose or glucose instead of sucrose and were
reported to be equally successful in the isolation of four Saccharomy-
ces species from oak samples in Portugal (Sampaio & Gonçalves,
2008) and the famous unknown parent of Saccharomyces pastorianus,
the cryotolerant yeast S. eubayanus (Gibson et al., 2017; Libkind et al.,
2011). Furthermore, not only have new species been described by
extensively sampling forests around the world but also these efforts
have augmented the number of strains available in S. cerevisiae (Q.
M. Wang, Liu, Liti, Wang, & Bai, 2012), S. paradoxus (Glushakova,
Ivannikova, Naumova, Chernov, & Naumov, 2007; Zhang, Skelton,
Gardner, & Goddard, 2010), Saccharomyces uvarum (Rodriguez,
Perez‐Traves, Sangorrin, Barrio, & Lopes, 2014), and S. eubayanus
(Peris et al., 2016; Rodriguez, Origone, Flores, & Lopez, 2016). Prelim-
inary identification of isolated yeasts can be accomplished with poly-
merase chain reaction amplification and sequencing of, for example,
the internal transcribed spacer region or the D1/D2 region of the
28S nuclear ribosomal large subunit rRNA gene (Schoch et al., 2012).
Metagenomic analysis of environmental samples after internal tran-
scribed spacer enrichment for Saccharomyces has been suggested as
a useful screening method for detected species that occur infre-
quently such as S. jurei (Alsammar et al., 2018). This approach could
also facilitate the discovery of new species. Altogether, these sampling
field trips have complemented our understanding of the ecology and
biodiversity of the Saccharomyces genus, as well as providing genetic
stocks for basic and applied sciences.
2.2 | Brewing properties of wild Saccharomyces
strains
The genetic diversity within the Saccharomyces genus can be exploited
for diverse fermentation processes (Cubillos, 2016; Steensels,
Meersman, et al., 2014; Steensels, Snoek, et al., 2014). In this context,
the brewing industry represents a field where new wild stocks or
hybrids can be potentially exploited to support the production of novel
flavours, which can be incorporated into craft beer during the fermen-
tation process (Table 1). For this, the phenotypic study of new wild Sac-
charomyces strains represents the first step towards determining
CUBILLOS ET AL.
whether the industrial potential of wild stocks is promising (Peter
et al., 2018; Warringer et al., 2011). Among the industrially relevant
traits of interest are ethanol and glycerol production, sugar utilization,
hop tolerance and low‐temperature resistance, and the relative pro-
duction of aroma compounds of interest, such as esters and higher
alcohols, as well as low levels of acetic acid and hydrogen sulphide
(Gallone et al., 2018; Gibson et al., 2017; Krogerus, Seppanen‐Laakso,
Castillo, & Gibson, 2017; Senkarcinova, Dias, Nespor, & Branyik,
2019; Steensels, Meersman, et al., 2014; Trindade de Carvalho et al.,
2017). The precise wort sugar profile depends on the raw ingredients
(mostly barley malt) and method of wort preparation, but the predom-
inant sugar sources in wort in decreasing order are maltose,
maltotriose, glucose, and fructose. Complete and timely conversion of
all sugars by Saccharomyces strains is a preferred outcome during the
process. In this context, not all species are able to consume the four
sugars. With the exception of S. cerevisiae, Saccharomyces species
appear to be unable to take up maltrotriose (Hebly et al., 2015;
Krogerus, Magalhães, Vidgren, & Gibson, 2015; Nguyen & Boekhout,
2017; Nikulin, Krogerus, & Gibson, 2018), thereby limiting their ability
to ferment wort efficiently. However, S. eubayanus seems highly profi-
cient in maltose utilization and exhibits other relevant traits appropriate
for brewing, including efficient biomass production at low temperature
(ideal for lager brewing), balancing the inability to consume maltrotriose
(Hebly et al., 2015; Krogerus et al., 2015; Mertens et al., 2015).
One of the coveted features of wild yeasts is their potential to
produce desirable volatile compounds for brewing (Mertens et al.,
2015; Nikulin et al., 2018). The type strain of S. arboricola, for example,
was seen to produce high levels of ethyl esters (Nikulin et al., 2018).
However, wild Saccharomyces isolates tend to produce significant per-
ceivable levels of phenolic off‐flavours (POF), mainly through the pro-
duction of 4‐vinyl guaiacol from ferulic acid (Coghe, Benoot, Delvaux,
Vanderhaegen, & Delvaux, 2004), imparting a clove‐like aroma to beer.
This is considered undesirable in most beer styles. The production of
4‐vinyl guaiacol is due to phenylacrylic acid decarboxylase and ferulic
acid decarboxylase, both interacting and playing a key step in detoxi-
fying phenolic acids (Richard, Viljanen, & Penttila, 2015). In some
cases, the POF aromas do not represent a problem but are instead
desired in the final fermented product, such as wheat beer, where
clove‐like aroma is part of the normal flavour profile (Gallone et al.,
2016). Nevertheless, the ability of wild yeast to ferment at low tem-
peratures, produce new aroma profiles, high ethanol levels, and low
H2S, and withstand stressful wort conditions represents potential for
new brewing strategies. Overall, these reports highlight that wild
yeasts can produce and contribute with alternative and/or new combi-
nations of tastes and aromas for beer. These could be used not only
for lager beer but also to innovate or accentuate aroma profiles in
other beer types. Commercial beers using wild stocks are already
available on the market and include beers produced with S. eubayanus
strains isolated from South America, North America, and Asia (Bing
et al., 2014; Libkind et al., 2011; Peris et al., 2014). The isolation of
a larger number of strains can expand the phenotypic diversity in each
species that we know of, providing a larger catalogue of isolates with
unique fermentation and aromatic profiles.
CUBILLOS ET AL. 387

TABLE 1 A selection of nonbrewing yeast recently evaluated for their brewing potential: Their source, isolation procedure (where reported), and
proposed application

Species Source Isolation procedure Application Reference

Arthroascus schoenii Insect, Mexico — Bioflavouring Gutiérrez et al., 2018
Barnettozyma Fruit, Russia — Bioflavouring Gutiérrez et al., 2018
californica
Brettanomyces spp. Beer, wine spoilage, soda — Bioflavouring, Holt et al., 2018
speciality beers
Candida humilis Dough, USA — Bioflavouring Gutiérrez et al., 2018
Cyberlindnera fabianii Fermented masau fruit, Malt extract agar with 0.1% Bioflavouring Nyanga et al., 2007;
Zimbabwe oxytetracycline and de Man, van Rijswijck et al., 2017
Rogosa, Sharpe agar with 0.1
natamycin
Cyberlindnera meyerae Plant, South Africa — Bioflavouring Gutiérrez et al., 2018
Galactomyces Soil, Puerto Rico Culture collection Bioflavouring Gutiérrez et al., 2018
geotrichum
Hanseniaspora vineae Ash bark, USA Ethanol‐supplemented rich media Sour beer Osburn et al., 2018
and hopped porter wort
Kazachstania servazzi Soil, Finland — Bioflavouring Gibson et al., 2015
Kluyveromyces lactis Dairy product — Bioflavouring Gutiérrez et al., 2018
Lachancea fermentati Birch bark, USA Ethanol‐supplemented rich media Sour beer Osburn et al., 2018
and hopped porter wort
Lachancea Wine, Italy; fruit, Czech Republic — Sour beer Domizio et al., 2016;
thermotolerans Gutiérrez et al., 2018
L. thermotolerans Shagbark hickory bark Ethanol‐supplemented rich media Sour beer Osburn et al., 2018
and hopped porter wort
Mrakia gelida Glacier meltwater, Italy — Low‐alcohol brewing De Francesco et al., 2018
Naumovozyma Salad, Finland — Bioflavouring Gibson et al., 2015
dairenensis
Pichia kudriavzevii Fermented masau fruit, Malt extract agar with 0.1% Bioflavouring Nyanga et al., 2007;
Zimbabwe oxytetracycline and de Man, van Rijswijck et al., 2017
Rogosa, Sharpe agar with 0.1
natamycin
Pichia kluyveri Commercial wine strain, Cacao — Bioflavouring Gutiérrez et al., 2018;
fermentation Holt et al., 2018
P. kluyveri Patent — Alcohol‐free beer Saerens & Swiegers, 2017
Pichia phlogasa Soil, South Africa — Bioflavouring Gutiérrez et al., 2018
Saccharomyces Cachaça distilleries, Brazil YP medium supplemented with 2% Lager and ale beer Araújo et al., 2018
cerevisiae sucrose, 0.1% chloramphenicol
S. cerevisiae Sourdough, Italy — Ale beer Marongiu et al., 2015;
Mascia et al., 2015
S. cerevisiae Brewing yeast starter culture, Wallerstein nutrient agar Ale beer Preiss et al., 2018
Norway
S. cerevisiae var Food supplement, Czech — Ale beer Senkarcinova et al., 2019
boulardii Republic
Saccharomyces Nothofagus spp., Argentina Yeast nitrogen base, raffinose Lager beer Gibson, Storgårds, Krogerus,
eubayanus & Vidgren, 2013; Libkind
et al., 2011
Saccharomycodes Patent — Alcohol‐free beer Haehn & Glaubitz, 1933;
ludwigii Huige et al., 1990
Scheffersomyces Patent — Alcohol‐free beer Li et al., 2011
shehatae
Schizosaccharomyces Oak bark, USA Ethanol‐supplemented rich media Sour beer Osburn et al., 2018
japonicus and hopped porter wort
Starmera caribaea Plant, West Indies — Bioflavouring Gutiérrez et al., 2018
Tetrapisispora blattae Insect — Bioflavouring Gutiérrez et al., 2018
Torulaspora delbrueckii Papaya leaf, Cameroon, beer, — Low‐alcohol beer, Canonico et al., 2016;
wine, brandy bioflavouring Michel et al., 2016

(Continues)
388 CUBILLOS ET AL.

TABLE 1 (Continued)

Species Source Isolation procedure Application Reference

Torulaspora Plant, USA — Bioflavouring Gutiérrez et al., 2018
microelliposoides
Wickerhamomyces Ragi, Indonesia — Bioflavouring Ravasio et al., 2018
anomalus
W. anomalus Shumard oak bark, USA Ethanol‐supplemented rich media Sour beer Osburn et al., 2018
and hopped porter wort
Wickerhamomyces Sugar — Bioflavouring Gutiérrez et al., 2018
subpelliculosus
Yarrowia lipolytica Fuel — Bioflavouring Gutiérrez et al., 2018
Zygosaccharomyces Kombucha, USA Pre‐screening in wort Alcohol‐free beer Bellut et al., 2018
bailii
Zygosaccharomyces Grape must, Italy; insect, — Alcohol‐free beer De Francesco et al., 2015;
rouxii Germany Gutiérrez et al., 2018

2.3 | Natural and artificial interspecies hybrids for
brewing
The utilization of wild Saccharomyces strains in brewing is not limited
to genetic monocultures; they can also serve as genetic stocks to gen-
erate new interspecies combinations that could exhibit hybrid vigour
for industrial traits (Hebly et al., 2015; Krogerus, Magalhães, Vidgren,
& Gibson, 2017; Mertens et al., 2015; Nikulin et al., 2018). In this con-
text, several natural Saccharomyces hybrids have been isolated from
industrial settings, such as wineries and beer fermentation tanks, dem-
onstrating that hybrid selection based on greater fitness under fer-
mentation conditions has already occurred. The best example is
S. pastorianus (S. cerevisiae × S. eubayanus) and reviewed elsewhere
(Gibson & Liti, 2015); however, many other interspecies hybrids have
also been recovered (Albertin et al., 2018; Gonzalez, Barrio, & Querol,
2008; Krogerus, Preiss, & Gibson, 2018; Peris et al., 2012). For exam-
ple, S. cerevisiae × Saccharomyces kudriavzevii hybrid strains were
recovered in brewing environments (Gonzalez et al., 2008; Peris
et al., 2012). That being said, natural hybrids are not found commonly
in nature compared with industrial settings. However, wild Saccharo-
myces strains with introgressions from other species do occur (Albertin
et al., 2018; Almeida et al., 2014; Peter et al., 2018)—some examples
are found in S. cerevisiae with introgressions from other species includ-
ing the closely related species S. paradoxus (Duan et al., 2018; Peter
et al., 2018). Still, most isolates exhibiting horizontal gene transfers
are generally associated with human activities: such as those found
in S. uvarum, where introgressions from S. eubayanus and S. kudriavzevii
were described (Albertin et al., 2018). These antecedents suggest that
hybrids are fitter solely when associated with fermentation environ-
ments but do not persist in the wild. Reduced fertility associated with
hybridization may be too great a sacrifice in natural systems where the
associated adaptability is required for survival (Gray & Goddard,
2012). Brewing environments in contrast are more consistent regard-
ing stresses encountered.
The greater number of isolates currently available allows design of
new artificial hybrids spanning the spectrum of desired phenotypes for
brewing. The isolation of S. eubayanus in Patagonia (Libkind et al.,
2011) represented the first step towards the generation of new
S. cerevisiae × S. eubayanus artificial hybrids for brewing, expanding
the limited genetic diversity of commercially available S. pastorianus
(Gibson et al., 2017). In this line, several reports explored the pheno-
typic profiles under brewing conditions of these artificial hybrids, in
terms of (a) aroma production, (b) fermentation capacity at low tem-
perature, and (c) maltose and maltotriose consumption (Krogerus
et al., 2015; Mertens et al., 2015; Nikulin et al., 2018). In general, these
reports clearly highlight that hybrids outperform parental strains dur-
ing the beer fermentation process and exhibit a broader temperature
tolerance. Interestingly, some of them showed industrial potential by
exhibiting production of fruit aromas, undetectable levels of POF, high
ethanol yields, and fast fermentation rates (Krogerus, Magalhães, et al.,
2017; Mertens et al., 2015). Currently, the number of new Saccharo-
myces strains, and particularly S. eubayanus isolates, is growing expo-
nentially, facilitating the generation and study of large numbers of
new intraspecies and interspecies hybrids covering a wider genetic
diversity within each species. Moreover, a grid of crosses can be gen-
erated between different species and strains to enrich the brewing
potential beyond S. pastorianus parental strains (Nikulin et al., 2018).
3 | ISOLATION AND BREWING POTENTIAL
O F N O N‐ S A C C H A R O M Y C E S SP P .
Saccharomyces is not, however, the only interesting genus from the per-
spective of brewing; beyond Saccharomyces, many yeasts with varying
characteristics can be found expanding the potential and the usability
of yeasts in brewing (Table 1). Usually, these species are isolated using
procedures suitable for a diverse range of species that possess traits
relevant to brewing and that preferably only select against unwanted
microorganisms. A common way to inhibit bacterial growth is to use
chloramphenicol (Laitila et al., 2006; Osburn, Ahmad, & Bochman,
2016) or by adjusting the conditions to make them less favourable to
bacteria, for example, by adding alcohol and/or lowering the pH of
the medium (Rosso, Lobry, Bajard, & Flandrois, 1995; Russell & Diez‐
Gonzalez, 1997). There is no single procedure applied universally for
this purpose but, rather, many different methods for different pur-
poses. A general approach is, however, to use enrichment media
CUBILLOS ET AL.
followed by selective agar plates for selection and isolation of single
colonies.
Ethanol tolerance is one widely sought‐after characteristic for
new brewing yeasts (Mukherjee et al., 2017; Osburn et al., 2016;
Ruyters et al., 2015). This can be favoured by using inhibitory ethanol
concentrations in one or more steps during the isolation procedure.
While looking for new yeast species suitable for brewing, Osburn
et al. (2016) and Osburn et al. (2018) used YPD8E5 medium (8% glu-
cose; 5% ethanol) or hopped porter wort (34 IBU with 5% of added
ethanol) for enrichment. Their survey yielded 284 strains in 54 differ-
ent species from a variety of substrates such as fruits, berries, and soils
but mainly from tree bark. The authors observed ethanol to be an effi-
cient inhibitor against fungal species as well, and most of their isolates
were ascomycetes. Others have reported similar findings: While
exploring the diversity of yeasts in wintering grapes, Sipiczki (2016)
noticed that high ethanol concentrations (>8% alcohol by volume
(ABV); 4‐week incubation) with raffinose as carbon source effectively
decreased the number of yeasts belonging to the Basidiomycota (from
69% of samples to 4%). From 270 mummified berries, he managed to
isolate 23 different ascomycete and 13 different basidiomycete yeast
species. Strains with high ethanol tolerance can also be isolated by
sampling from high alcohol environments, as demonstrated by
Choudhary, Singh, and Nain (2017) who isolated yeasts for bioethanol
production from distillery wastes. Indeed, both of their distillery iso-
lates, Wickerhamomyces anomalus and S. cerevisiae, were good pro-
ducers of ethanol. As demonstrated by these studies, alcohol can be
used as an efficient tool in isolation procedures to select for isolates
of species already associated with fermentations. However, the use
of alcohol might inhibit the growth of yeasts that could be suitable
for low‐ or non‐alcoholic beer production, and thus, alcohol‐free pro-
cedures should be considered as well. Laitila et al. (2006) mixed their
samples with saline water, homogenized them, and plated diluted sam-
ples directly on yeast–malt extract agars, which contained antibiotics
and Triton X but no alcohol. Their survey of yeasts in malting yielded
28 ascomycete species and 18 basidiomycete species.
Wort is one of the simplest selective media for isolating yeasts. It
favours species that are already capable of utilizing the sugars and
nutrients available in the brewer's wort and can be adjusted to min-
imize bacterial contamination simply by lowering the pH and thereby
creating less hospitable conditions for many bacterial species but
without limiting the growth of the more acid‐resistant yeast species,
and/or adding ethanol, typically to concentrations of 3–5% ABV
(Rosso et al., 1995; Russell & Diez‐Gonzalez, 1997). After this, it
can be used as a medium to inoculate fruits and berries, or it can
be left uncovered to collect microbes directly from air. This method
is popular among home brewers, who are attempting to isolate their
own wild yeasts (e.g., http://www.milkthefunk.com/wiki/Wild_Yeast_
Isolation). Anecdotally, the method appears effective; though, limited
peer‐reviewed studies have been published. Spontaneous wort fer-
mentations have been shown to exhibit complex population dynam-
ics, and the microflora varies during the cultivation period (Spitaels
et al., 2014). In order to fully exploit the available diversity, one
may need to sample repeatedly to obtain isolates representative of
389
the various fermentation stages. It may also mask some of the
species in sympatric relationship by favouring one over the other
(see Sampaio & Gonçalves, 2008 for the isolation of different
Saccharomyces spp.).
3.1 | Brewing properties
As a diverse group of fungi, the phenotypic properties of wild yeasts
vary greatly. It is also noteworthy that strains within a species may vary
considerably and that the sought after functionality may not be found in
every isolate of the species (Domizio et al., 2016; Michel et al., 2016;
Osburn et al., 2018) highlighting the importance of a careful screening.
Recent studies have revealed yeast species capable of producing
lactic acid in beer fermentations. One such yeast is L. thermotolerans
—the ability of which to produce acids was first reported in studies
on wine (Kapsopoulou, Kapaklis, & Spyropoulos, 2005; Mora, Barbas,
& Mulet, 1990). It was later found that this species was able to simul-
taneously ferment brewer's wort while producing appreciable levels of
lactic acid as well, making it possible to produce sour beer without
added bacteria (Domizio et al., 2016). This yeast seems to have a
widespread distribution but appears to be frequently associated with
insects such as yellow jacket wasps and fruit flies as well as oak trees
(Babcock et al., 2018; Hranilovic et al., 2018). Osburn et al. (2018) iso-
lated strains of Lachancea fermentati, Hanseniaspora vineae,
W. anomalus, and Schizosaccharomyces japonicus exhibiting similar
properties. Beer pH levels varied between 3.20 (S. japonicus) and
3.74 (L. fermentati). The highest attenuation rate (of 83%) was
reported with W. anomalus suggesting, unusually, that it was capable
of using maltotriose, whereas the rest of the species reached values
between 40% and 70% in laboratory fermentations.
Another potential use for wild yeasts is bioflavouring, where the
fermentation rates of the wild yeasts are secondary, and the focus
rather relies on increased production of desirable and often unique
aroma compounds. Here, fermentations are often carried out as
cofermentations, where the alcohol production is achieved with, for
example, ale or lager strains. Gibson et al. (2015) reported the use of
Kazachstania servazzi and Naumovozyma dairenensis for enhancing
the aroma profiles of lager beer (levels of 3‐methylbutanol,
phenylethanol, and phenylethyl acetate as well as ethyl decanoate
and caprylate were increased). Holt et al. (2018) found a strain of
Pichia kluyveri with a high production of isoamyl acetate (over
80 mg/L after 2 days of fermentation) as an interesting candidate for
sequential fermentation. T. delbrueckii strains have been proposed as
bioflavouring agents, with significant changes, both increases and
decreases, seen in the concentration of individual volatile flavour com-
pounds when the yeasts are involved in cofermentation with produc-
tion strains (Canonico, Agarbati, Comitini, & Ciani, 2016; Michel et al.,
2016). Brettanomyces species were, however, shown to be able to pro-
duce 4‐ethyl guaiacol (spicy, vanilla flavour) in sequential fermenta-
tions. In the study of Ravasio et al. (2018), W. anomalus was used
successfully for enhancing the aroma profile of lager beer in
cofermentation with a regular lager yeast. Cofermentations with
390 CUBILLOS ET AL.

Cyberlindnera fabianii or Pichia kudriavzevii have likewise resulted in

greater aroma content relative to alcohol level. In the case of the
C. fabianii cocultures, the ratio of brewer's yeast cells to C. fabianii cells
at inoculation had a clear impact on beer composition, thereby
affording the brewer some control over the quality of the final product
(van Rijswijck, Wolkers‐Rooijackers, Abee, & Smid, 2017).
In the production of low‐ and non‐alcoholic beers, inability to
reach high attenuation rates is a desirable characteristic (Gibson
et al., 2017). One such species is Saccharomycodes ludwigii, which
has a long history in the production of non‐alcoholic beers (Haehn &
Glaubitz, 1933; Huige, Sanchez, & Leidig, 1990). New strains are
sought, but in addition to low alcohol productivity, suitable strains
need to fulfil certain requirements, that is, a good portion of pleasant
aroma compounds and low levels of undesirable flavours—which
makes the task more challenging (De Francesco, Turchetti, Sileoni,
Marconi, & Perretti, 2015). Other species mentioned in patented
methods for the production of low‐ and non‐alcoholic beers are
Scheffersomyces shehatae (formerly Candida shehatae; Li, Liu, & Zhang,
2011) and P. kluyveri (Saerens & Swiegers, 2017). Strains of
Zygosaccharomyces rouxii have also been studied for production of
low‐alcohol beers, though the strains had ability to produce alcohol
over 1.0% ABV (De Francesco et al., 2015). The basidiomycete yeast
M. gelida has been found to produce low‐alcohol beers with a flavour
superior to that of S. ludwigii (De Francesco et al., 2018). The M. gelida
strain was isolated from glacier meltwater and was characterized by
strong resistance to low temperatures. Strains of this species typically
do not grow at temperatures of 17°C and above, a characteristic that
may facilitate application at lager‐brewing temperatures.

4 | R E P U R P O S I N G OF Y E A S T S F R O M
DIFFERENT FOOD AND BEVERAGES

4.1 | Traditional fermented beverages

Traditional fermented beverages may also be an important source of

new yeasts for the brewing industry. Many traditionally fermented
beers and other beverages are initiated spontaneously and typically
contain a mixture of native yeasts in addition to S. cerevisiae. Collec-
tively, the yeasts are responsible for the conversion of fermentable
sugars into ethanol as well as the production of a high number of
flavour‐active compounds, many of which are beverage specific
(Varela, 2016). Yeasts found naturally in these substrates represent a
FIGURE 2 Yeasts with potential for use in industrial fermentation
potentially valuable source of production strains with beneficial char- have been isolated from traditional fermented beverages such as (a)
acteristics such as enhanced resistance, or new flavour and aroma pro- sourdough cultures for baking, (b) chicha from Latin America, and (c)
files that could be applied in a controlled way to industrial brewery traditional farmhouse ales (shown is a yeast ring used for maintenance
systems (Faria‐Oliveira et al., 2015; Figure 2). of Norwegian kveik yeast in dried form). Images are supplied by Nikola
Michael, Blanca Vallejos, and Lars Marius Garshol, respectively [Colour
Beer production from barley malt is currently the most common
figure can be viewed at wileyonlinelibrary.com]
brewing process worldwide. Nevertheless, there is a wide diversity
of traditional beers that have been produced around the world using
different types of starchy plants such as maize (in South America), 2012). The existence of a microbial consortium including bacteria
soy (in India and Iran), millet and sorghum (in Africa), and rice (in the and yeasts is typical in these fermentations (Blandino, Al‐Aseeri,
Far East), most of them made in an artisanal way (Marshall & Mejia, Pandiella, Cantero, & Webb, 2003). Currently, there are several
CUBILLOS ET AL.
publications related to the study of yeasts with fermentative potential
isolated from traditional beverages; these include traditional beers of
South America (Mendoza, Neef, Vignolo, & Belloch, 2017; Piló et al.,
2018; Vallejo et al., 2013), Africa (Jespersen, 2003; Lyumugabe, Gros,
Nzungize, Bajyana, & Thonart, 2012), Asia (Bing et al., 2014), and
Europe (Spitaels et al., 2015), with Saccharomyces strains being the
most studied. There is a considerable phenotypic diversity among
S. cerevisiae strains isolated from various environments, due to large
fluctuations in gene expression levels and environmental stress selec-
tion (Dussap & Poughon, 2017). Natural Saccharomyces strains iso-
lated from Peruvian chichas had a phenotypic profile that differed
signficantly compared with standard production strains. In particular,
they had a unique capacity to ferment trehalose and to asimilate var-
ious carbon sources such as glucosamine, arabinose, melibiose, cellobi-
ose, and starch (Vallejo et al., 2013). Melibiose and trehalose
assimilation are not common characteristics of ale strains and could
even be criteria for differentiation (Berlowska, Kregiel, & Rajkowska,
2015). A range of S. cerevisiae strains, genetically distinct from other
brewing yeasts, and a S. cerevisiae × S. uvarum hybrid were recently
isolated from traditional Norwegian farmhouse brewing yeast cultures,
known as kveik (Krogerus, Preiss, et al., 2018; Preiss et al., 2018). The
isolates were characterized by multiple brewing‐relevant properties,
such as high flocculation potential, high ethanol tolerance, high tem-
perature tolerance, absence of phenolic flavour production, and high
rates of fermentation in wort.
There is a growing interest in the study of new yeasts isolated from
nonbrewing environments that can be co‐opted for beer production.
Traditional fermented beverages, other than beers, can be a rich source
of new fermentative yeasts. Species involved in spontaneous fermenta-
tions destined for distilled products, such as cachaça, tequila, mezcal,
and sake, among others, are an important source of new yeasts for alco-
holic beverages (Gschaedler, 2017). Beverages produced with these
yeasts can exhibit unique chemical compositions and organoleptic char-
acteristics. In some cases, there is also a significant impact on the nutri-
tional value of the final product, which contributes to quality
improvement and diversification. Takao et al. (2018) showed that sake
brewed with an indigenous yeast of sake brewery (Km67) had fuller
body and unique sensory properties (stronger saltiness, sourness, bitter-
ness, and astringency) without compromising ethanol production. Addi-
tionally, Km67 had higher stress tolerance related with genetic
differences to the industrial K7 yeast. It has also been shown that a
higher yeast population in pulque, a Mexican traditional beverage, is
related with a higher nutritional value related to its content of vitamins
(B complex and vitamin C), essential amino acids (lysine and tryptophan),
and available iron (Escalante et al., 2016; Lappe‐Oliveras et al., 2008).
Moreover, the use of native strains in tequila production (S. cerevisiae,
Kluyveromyces marxianus, and Starmerella magnoliae) is associated with
a higher amount of volatile compounds (terpenes and isoamyl and
isobutyl alcohol), suggesting that the different enzymatic and metabolic
activities of each strain play an important role in the aromatic profile of
the final beverage (Lappe‐Oliveras et al., 2008). S. cerevisiae strains iso-
lated from cachaça, for example, have presented interesting traits for
beer and bioethanol production. Three wild‐type strains of S. cerevisiae
391
from Brazilian cachaça distilleries have shown good performance in
terms of ethanol productivity and yield and a lower accumulated cell
damage during and after two rounds of fermentation (Paredes et al.,
2018). Araújo et al. (2018) isolated two strains from cachaça distilleries
that had suitable characteristics to produce lager and ale beer styles,
such as capacity to ferment maltose and maltotriose, ability to grow at
low temperatures, low H2S production, and appropriate flocculation
profile. Furthermore, a new diploid hybrid yeast strain generated by
direct mating with two native S. cerevisiae yeast strains from cachaça
distilleries showed interesting characteristics for lager beer production
(low temperature fermentation, high flocculation capacity, and desirable
acetate esters production); nevertheless, POF character and their
inability to ferment maltotriose should be also considered (Figueiredo
et al., 2017).
Traditional fermented beverages could also be an important
source of new strains of non‐Saccharomyces yeasts for the production
of alcohol‐free beers. Bellut et al. (2018) showed the highest potential
of a Zygosaccharomyces bailii strain, isolated from Kombucha, in the
brewing of alcohol‐free beer. In addition to its inability to ferment
maltose and maltotriose, it was able to grow at high iso‐α‐acid con-
centrations, was POF negative, and was moderately flocculent, and
the final beer was slightly fruitier and had low level of worty charac-
ter. Candida tropicalis, isolated from traditionally fermented beverages
from African countries, was studied as an alternative for sorghum
beer production and low ethanol concentration, and few aroma com-
pounds were reported (N'Guessan, N'Dri, Camara, & Djè, 2010).
Pichia spp. have been isolated from different traditional beverages
from Latin America. Many of these species have shown high ester
production (isoamyl acetate and ethyl acetate) in the fermentation
of agave juice (Ramos & Schwan, 2017; Segura‐García, Taillandier,
Brandam, & Gschaedler, 2015). A recent patent used P. kluyveri to
produce a low‐alcohol beer (Saerens & Swiegers, 2017); new isolates
of this species from traditional beverages could likewise be applied
in breweries.
4.2 | Baking yeasts
Of the various food systems from which new brewing yeasts may be
derived, baking seems to be one of the promising. In both brewing
and baking, yeast is required to ferment cereal‐derived sugars, with
the fermentative end products, ethanol in the case of beer, and carbon
dioxide in the case of bread, being essential for the production pro-
cess. It is not surprising that there exist phenotypic similarities
between the two groups. Both are capable of maltose, and often
maltotriose, utilization, and copy numbers for genes related to the
uptake and assimilation of these sugars are over‐represented in both
groups, as are genes related to osmotolerance (Gonçalves et al.,
2016). The similarities observed are not necessarily a case of conver-
gent evolution in similar substrates. Historically, the same yeast popu-
lations have often been used for both brewing and baking. Writing in
the 1st century AD, Pliny the Elder describes the use of yeast
skimmed from brewing vessels for baking (Bostock & Riley, 1855).
392
Also, in ancient Egypt, the brewing process involved first lightly baking
bread from emmer wheat and/or barley. This bread would then be
broken and strained with water through a sieve before being
fermented, presumably with the same yeast that carried out the bak-
ing (Lutz, 1922). Use of the same yeast for both processes still occurs
today; the bread‐based Kvass beer is still brewed in the Baltic states,
Eastern Europe, and Russia (Loponen & Sibakov, 2013), and traditional
Nordic beers such as Finland's sahti are often brewed using baker's
yeast rather than brewer's yeast (Ekberg et al., 2013). The historical
multifunctionality of yeast is reflected in the genomes of both groups.
Comparisons of industrial yeast genomes have repeatedly shown a
close relatedness of brewing and baking yeasts compared with other
groups (Gallone et al., 2016, Gonçalves et al., 2016; Legras et al.,
2018; Legras et al., 2007; Peter et al., 2018).
Baking yeasts may therefore represent a useful source of new
brewing yeast strains. Indeed, in brewing trials carried out with yeasts
sourced from different food and beverage systems (wine, cider, and bak-
ing), it was the baking yeast that showed most potential for brewing
(Rossi, Turchetti, Sileoni, Marconi, & Perretti, 2018). Marongiu et al.
(2015) found that many S. cerevisiae strains isolated from sourdoughs
were able to efficiently ferment wort, and pilot‐scale fermentations pro-
duced beer of the same quality as that produced by a commercial
brewing yeast culture. This result was confirmed by Mascia et al. (2015)
and Mascia, Fadda, Kerabín, Dostálek, and Del Caro (2016), who also
compared sourdough isolates with a commercial culture, in this case for
wheat beer brewing. Beers fermented with the sourdough‐derived cul-
tures were found to be equivalent to or better than a commercial brewing
strain with regard to yield and flavour profile. Direct application of sour-
dough liquid culture to wort, without any prior attempt to isolate individ-
ual strains, was also performed in an effort to produce acidic beers with
complex flavour profiles similar to Belgian lambic beers (Ripari,
Tomassetti, Cecchi, & Enrico, 2018). Though apparently successful, this
approach may have limited utility outside of small‐scale craft brewing.
Brewer's yeast is characterized by strong flocculation and lack of 4‐
vinylguaiacol production (Mukai, Masaki, Fujii, Kawamukai, & Iefuji,
2010; Vidgren & Londesborough, 2011). These traits may not be
expected in baker's yeasts, and the available reports from studies involv-
ing baking yeasts in brewing have not mentioned these features. Further
characterization is clearly required, and it may be expected that yeasts
isolated from bread will produce phenolic flavour in beer and may also
require process steps for beer clarification (Gallone et al., 2016).
Sourdoughs are also a potentially useful source of
non‐Saccharomyces ascomycete yeasts for production of low‐alcohol
beers (see above). Such yeasts benefit from the maltose‐hydrolysing
activities of sourdough bacteria such as Lactobacillus sanfranciscensis
and require only the ability to take up monomeric sugars.
Non‐Saccharomyces yeasts resident in sourdoughs include species
such as T. delbrueckii or W. anomalus that have Generally Recognized
as Safe/Qualified Presumption of Safety status (De Vuyst, Harth,
Van Kerrebroeck, & Leroy, 2016; De Vuyst et al., 2014). Also, such
yeasts, because they are isolated from natural food sources, may be
considered more acceptable by brewers and consumers when com-
pared with yeast isolated from nonfood sources. When considering
CUBILLOS ET AL.
sourdough cultures as a potential source of yeast for brewing, a num-
ber of factors must be considered. First, sourdough cultures often host
a limited number of species, with approximately six different species
found frequently. Furthermore, one or two species of yeast typically
dominate individual cultures, thereby limiting the potential for
bioprospecting (De Vuyst et al., 2014). The yeast species associated
with sourdough appear to be relatively consistent, with differences in
assemblages usually due to differences in processes and raw materials
associated with regions rather than the regions per se. Metschnikowia
pulcherrima isolated from a southern Italian sourdough, for example,
was linked to fruit used in the process rather than being associated with
the microbiota of the region (Palomba, Blaiotta, Ventorino, Saccone, &
Pepe, 2011). Also, yeasts found in laboratory‐generated sourdoughs
(where flour is the only source of yeasts) will be different to those found
in sourdoughs associated with bakeries (where provenance can vary).
Although S. cerevisiae is typical of both laboratory and bakery sour-
doughs, it appears to be more abundant in the latter. In contrast, Candida
glabrata and W. anomalus are more frequently isolated from the labora-
tory cultures (Vrancken et al., 2010). The grain used also appears to influ-
ence the microbes present with, for example, oat and buckwheat not
apparently supporting yeast growth (Moroni, Arendt, & Dal Bello,
2011), whereas P. kudriavzevii is associated with nonwheat sourdoughs
(Vogelmann & Hertel, 2011).
One of the main benefits of sourdoughs from the bioprospector's
perspective is the ease of isolating the abundant yeast cells and the
general absence of mycelial fungal contaminants present due to the
antifungal properties of lactic acid bacteria (LAB) (Hassan &
Bullerman, 2008). Bacteria are readily removed during isolation by
the use of antibiotics such as chloramphenicol (Pulvirenti, Solieri,
Gullo, De Vero, & Giudici, 2004). Overall, sourdoughs appear to rep-
resent a reliable source of abundant and safe yeasts that can be uti-
lized in brewing. Other solid‐state fermentation systems such as
cacao, cheese, and coffee fermentations may similarly act as a rich
source of yeasts that could be utilized in brewing. This has at least
been observed for P. kluyveri isolates obtained from cacao fermenta-
tion (Holt et al., 2018) and a dairy isolate of Kluyveromyces lactis
(Gutiérrez, Boekhout, Gojkovic, & Katz, 2018).
5 | PERSPECTIVES
Conventional brewing yeasts have evolved through processes of nat-
ural and artificial selection to efficiently ferment high‐quality beer.
However, yeast strains in use in brewing today presumably represent
only a subset of the strains available before beer production became
industrialized and centralized. Furthermore, currently used brewing
yeasts have a specific phenotypic range that limits the potential for
truly novel flavour profiles. As we have highlighted above, there is
considerable potential for exploiting the brewing properties of non-
conventional yeast to produce unconventional beers. A case in point
is the production of flavoursome low‐alcohol beers by maltose‐
negative, non‐Saccharomyces yeasts to meet the demands of a chang-
ing market (Gutiérrez et al., 2018).
CUBILLOS ET AL. 393

Yeasts that have evolved in natural or nonbrewing environments TABLE 2 The ecological origin of the 159 strains out of 1,011
may not possess typical signatures of domestication such as strong described by Peter et al. (2018) carrying homozygous nonsense or
flocculation, maltotriose utilization, or absence of POF character. frameshift mutations on PAD1 or FDC1

These issues are however surmountable. First, the modern brewing Ecological origin Number of strains
industry has at its disposal process steps to overcome these chal- Bakery 8
lenges. Poor flocculation is a feature of many brewing yeasts, and Beer 28
industrial centrifuges are available for beer clarification. In addition, Cider 1
mashing conditions may be optimized to reduce the proportion of fer- Distillery 5
mentable sugar consisting of maltotriose or the level of ferulic acid Fermentation 9
(the precursor of the 4‐vinylguiacol off‐flavour). Second, wild yeast Flower 1
traits may be reduced or removed through evolutionary engineering Fruit 3
and selection. Strains of POF‐negative S. eubayanus strains have, for Human, clinical 6
example, been created for brewing applications (Diderich, Weening, Industrial 3
van den Broek, Pronk, & Daran, 2018), maltotriose utilization may be Lab strain 1
increased (Brickwedde et al., 2017), and ethanol tolerance can be Nature 7
enhanced (Krogerus, Holmström, & Gibson, 2018). The adaptability Palm wine 17
of yeasts is evident in their genomes. Some yeast strains used for Sake 46
brewing do not in fact belong to any brewing yeast clade. Wine yeast Soil 3
strains have been employed to brew beer and vice versa, whereas Tree 3
S. cerevisiae ale yeast has also been employed for lager brewing Unknown 3
(Gallone et al., 2016). Wine yeast has also been discovered in cachaça Water 6
fermentations, where evidence of secondary domestication is evident Wine 9
(Barbosa et al., 2018).
As the number of published yeast genome sequences rapidly
predoctoral grant awarded. The Academy of Finland (Academy
increases, an opportunity also arises for identifying in silico unique
Projects 276480 and 305453) is gratefully acknowledged for
strains suitable for brewing. By using knowledge on the genetic loci
supporting the work of B. Gibson. J. Nikulin is supported by Fonds
associated with various relevant traits, it is possible to rapidly screen
Baillet Latour. K. Krogerus wishes to thank the Alfred Kordelin Foun-
thousands of strains. For brewing applications, this could involve
dation, Svenska Kulturfonden—The Swedish Cultural Foundation in
looking for the presence of genes encoding functional maltose and
Finland, and Suomen Kulttuurirahasto for their support. Nikola
maltotriose transporters or mutations in genes associated with the
Michael, Blanca Vallejos, and Lars Marius Garshol are thanked for pro-
synthesis of aroma compounds (Gallone et al., 2016; Gonçalves
viding images of, respectively, the sourdough culture, chicha, and
et al., 2016). To demonstrate, we screened the publically available
kveik yeast ring in Figure 2.
sequence data (obtained from https://www.yeastgenome.org/1011‐
yeast‐genomes) of the 1,011 S. cerevisiae strains described by Peter
et al. (2018) for homozygous nonsense or frameshift mutations in ORCID
PAD1 and FDC1 genes. When functional, these genes are responsible
Francisco A. Cubillos https://orcid.org/0000-0003-3022-469X
for the formation of POF, which are undesirable in many beer styles
Brian Gibson https://orcid.org/0000-0002-1270-6532
(Mukai et al., 2010). A total of 159 potentially POF‐negative strains
Kristoffer Krogerus https://orcid.org/0000-0001-7694-8277
were found, and 131 of these were isolated from substrates other
than beer (Table 2). Furthermore, 138 of these strains were not RE FE RE NC ES
assigned to the “African beer,” “ale beer,” or “Mosaic beer” clades. Albertin, W., Chernova, M., Durrens, P., Guichoux, E., Sherman, D. J.,
Such in silico screens could therefore be useful for identifying Masneuf‐Pomarede, I., & Marullo, P. (2018). Many interspecific chro-
brewing‐relevant strains from previously isolated and sequenced mosomal introgressions are highly prevalent in Holarctic
Saccharomyces uvarum strains found in human‐related fermentations.
strains and also save time and resources that would otherwise be
Yeast, 35, 141–156. https://doi.org/10.1002/yea.3248
needed for laboratory‐based screenings. There is also the possibility
Almeida, P., Gonçalves, C., Teixeira, S., Libkind, D., Bontrager, M., Masneuf‐
of applying this strategy in the pursuit of wild or nonconventional Pomarede, I., … Sampaio, J. (2014). A Gondwanan imprint on global
yeast strains possessing brewing‐relevant characteristics (Viigand, diversity and domestication of wine and cider yeast Saccharomyces
Põšnograveja, Visnapuu, & Alamäe, 2018). uvarum. Nature Communications, 5, 4044. https://doi.org/10.1038/
ncomms5044
Alsammar, H. F., Naseeb, S., Brancia, L. B., Gilman, R. T., Wang, P., &
ACKNOWLEDGEMEN TS Delneri, D. (2018). Targeted metagenomics approach to capture
the biodiversity of Saccharomyces genus in wild environments.
N. Grijalva Vallejos acknowledges the “Secretaría de Educación Supe- Environmental Microbiology Reports. https://doi.org/10.1111/1758‐22
rior, Ciencia, Tecnología e Innovación” (SENESCYT) of Ecuador, for the 29.12724
394
Aquilani, B., Laureti, T., Poponi, S., & Secondi, L. (2015). Beer choice and
consumption determinants when craft beers are tasted: An exploratory
study of consumer preferences. Food Quality and Preference, 41,
214–224. https://doi.org/10.1016/j.foodqual.2014.12.005
Araújo, T. M., Souza, M. T., Diniz, R. H. S., Yamakawa, C. K., Soares, L. B.,
Lenczak, J. L., … Brandão, R. L. (2018). Cachaça yeast strains: Alternative
starters to produce beer and bioethanol. Antonie Van Leeuwenhoek, 111,
1749–1766. https://doi.org/10.1007/s10482‐018‐1063‐3
Babcock, T., Borden, J., Gries, R., Carroll, C., Moore, M., & Gries, G. (2018).
Lachancea thermotolerans, enhances attraction of three yellowjacket
species (Hymenoptera: Vespidae) to fruit powder. Environmental Ento-
mology, nvy139, 47(6), 1553–1559.
Barbosa, R., Pontes, A., Santos, R. O., Montandon, G., de Ponzzes‐Gomes,
C., Morais, P., … Sampaio, J. P. (2018). Multiple rounds of artificial selec-
tion promote microbe secondary domestication. Genome Biology and
Evolution, 10(8), 1939–1955. https://doi.org/10.1093/gbe/evy132
Barnett, J. A., & Lichtenthaler, F. W. (2001). A history of research on yeasts
3: Emil Fischer, Eduard Buchner and their contemporaries, 1880–1900.
Yeast, 18, 363–388. https://doi.org/10.1002/1097‐0061(200103
15)18:4<363::AID‐YEA677>3.0.CO;2‐R
Bellut, K., Michel, M., Zarnkow, M., Hutzler, M., Jacob, F., De Schutter, D.,
… Arendt, E. (2018). Application of non‐Saccharomyces yeasts isolated
from Kombucha in the production of alcohol‐free beer. Fermentation,
4, 66. https://doi.org/10.3390/fermentation4030066
Berlowska, J., Kregiel, D., & Rajkowska, K. (2015). Biodiversity of brewery
yeast strains and their fermentative activities. Yeast, 32, 289–300.
https://doi.org/10.1002/yea.3041
Bing, J., Han, P. J., Liu, W. Q., Wang, Q. M., & Bai, F. Y. (2014). Evidence for
a Far East Asian origin of lager beer yeast. Current Biology, 24(10),
R380–R381. https://doi.org/10.1016/j.cub.2014.04.031
Blandino, A., Al‐Aseeri, M. E., Pandiella, S. S., Cantero, D., & Webb, C. (2003).
Cereal‐based fermented foods and beverages. Food Research International,
36(6), 527–543. https://doi.org/10.1016/S0963‐9969(03)00009‐7
Bostock, J., & Riley, H. T. (1855). The natural history of Pliny vol 1. London,
UK: Henry Bohn.
Brickwedde, A., van den Broek, M., Geertman, J.‐M. A., Magalhães, F.,
Kuijpers, N. G. A., Gibson, B., … Daran, J.‐M. G. (2017). Evolutionary engi-
neering in chemostat cultures for improved maltotriose fermentation
kinetics in Saccharomyces pastorianus lager brewing yeast. Frontiers in
Microbiology, 8, 1690. https://doi.org/10.3389/fmicb.2017.01690
Canonico, L., Agarbati, A., Comitini, F., & Ciani, M. (2016). Torulaspora
delbrueckii in the brewing process: A new approach to enhance
bioflavour and to reduce ethanol content. Food Microbiology, 56,
45–51. https://doi.org/10.1016/j.fm.2015.12.005
Choudhary, J., Singh, S., & Nain, L. (2017). Bioprospecting thermotolerant
ethanologenic yeasts for simultaneous saccharification and fermentation
from diverse environments. Journal of Bioscience and Bioengineering,
123(3), 342–346. https://doi.org/10.1016/j.jbiosc.2016.10.007
Coghe, S., Benoot, K., Delvaux, F., Vanderhaegen, B., & Delvaux, F. R.
(2004). Ferulic acid release and 4‐vinylguaiacol formation during
brewing and fermentation: Indications for feruloyl esterase activity in
Saccharomyces cerevisiae. Journal of Agricultural and Food Chemistry,
52, 602–608. https://doi.org/10.1021/jf0346556
Crauwels, S., Van Assche, V., de Jonge, R., Borneman, A. R., Verreth, C., Troels,
P., … Lievens, B. (2015). Comparative phenomics and targeted use of geno-
mics reveals variation in carbon and nitrogen assimilation among different
Brettanomyces bruxellensis strains. Applied Microbiology and Biotechnology,
99, 9123–9134. https://doi.org/10.1007/s00253‐015‐6769‐9
Cubillos, F. A. (2016). Exploiting budding yeast natural variation for indus-
trial processes. Current Genetics, 62, 745–751. https://doi.org/
10.1007/s00294‐016‐0602‐6
CUBILLOS ET AL.
De Francesco, G., Sannino, C., Sileoni, V., Marconi, O., Filippucci, S.,
Tasselli, G., & Turchetti, B. (2018). Mrakia gelida in brewing process:
An innovative production of low alcohol beer using a psychrophilic
yeast strain. Food Microbiology, 76, 354–362. https://doi.org/
10.1016/j.fm.2018.06.018
De Francesco, G., Turchetti, B., Sileoni, V., Marconi, O., & Perretti, G.
(2015). Screening of new strains of Saccharomycodes ludwigii and
Zygosaccharomyces rouxii to produce low‐alcohol beer. Journal of the
Institute of Brewing, 121(1), 113–121. https://doi.org/10.1002/jib.185
De Vuyst, L., Harth, H., Van Kerrebroeck, S., & Leroy, F. (2016). Yeast
diversity of sourdoughs and associated metabolic properties and func-
tionalities. International Journal of Food Microbiology, 239, 26–34.
https://doi.org/10.1016/j.ijfoodmicro.2016.07.018
De Vuyst, L., Van Kerrebroeck, S., Harth, H., Huys, G., Daniel, H.‐M., &
Weckx, S. (2014). Microbial ecology of sourdough fermentations:
Diverse or uniform? Food Microbiology, 37, 11–29. https://doi.org/
10.1016/j.fm.2013.06.002
Diderich, J., Weening, J., van den Broek, M., Pronk, J., & Daran, J. (2018).
Selection of Pof‐Saccharomyces eubayanus variants for the construc-
tion of S. cerevisiae × S. eubayanus hybrids with reduced 4‐vinyl
guaiacol formation. Frontiers in Microbiology, 9, 1640.
Domizio, P., House, J. F., Joseph, C. M. L., Bisson, L. F., & Bamforth, C. W.
(2016). Lachancea thermotolerans as an alternative yeast for the pro-
duction of beer. Journal of Institute of Brewing, 122, 599–604.
https://doi.org/10.1002/jib.362
Duan, S.‐F., Han, P.‐J., Wang, Q.‐M., Liu, W.‐Q., Shi, J.‐Y., Li, K., … Bai, F.‐Y.
(2018). The origin and adaptive evolution of domesticated populations
of yeast from Far East Asia. Nature Communications, 9, 2690. https://
doi.org/10.1038/s41467‐018‐05106‐7
Dussap, C. G., & Poughon, L. (2017). Microbiology of alcoholic fermenta-
tion. In A. Pandey, M. A. Sanromán, G. Du, C. R. Soccol, & C.‐G.
Dussap (Eds.), Current developments in biotechnology and bioengineering
(pp. 263–269). London, UK: Elsevier. https://doi.org/10.1016/B978‐0‐
444‐63666‐9.00010‐8
Ekberg, J., Rautio, J., Mattinen, L., Vidgren, V., Londesborough, J., &
Gibson, B. R. (2013). Adaptive evolution of the lager brewing yeast
Saccharomyces pastorianus for improved growth under hyperosmotic
conditions and its influence on fermentation performance.
FEMS Yeast Research, 13, 335–349. https://doi.org/10.1111/1567‐
1364.12038
Escalante, A., López Soto, D. R., Velázquez Gutiérrez, J. E., Giles‐Gómez,
M., Bolívar, F., & López‐Munguía, A. (2016). Pulque, a traditional Mex-
ican alcoholic fermented beverage: Historical, microbiological, and
technical aspects. Frontiers in Microbiology, 7, 1026.
Faria‐Oliveira, F., Diniz, R. H. S., Godoy‐Santos, F., Piló, F. B., Mezadri, H.,
Castro, I. M., & Brandão, R. L. (2015). The role of yeast and lactic acid
bacteria in the production of fermented beverages in South America.
In Food production and industry. IntechOpen, London, UK: InTech.
https://doi.org/10.5772/60877
Figueiredo, B. I. C., Saraiva, M. A. F., de Souza Pimenta, P. P., de Souza
Testasicca, M. C., Sampaio, G. M. S., da Cunha, A. C., … Brandão, R. L.
(2017). New lager brewery strains obtained by crossing techniques
using cachaça (Brazilian spirit) yeasts. Applied and Environmental Micro-
biology, 83(20), e01582‐17. https://doi.org/10.1128/AEM.01582‐17
Gallone, B., Mertens, S., Gordon, J. L., Maere, S., Verstrepen, K. J., &
Steensels, J. (2018). Origins, evolution, domestication and diversity of
Saccharomyces beer yeasts. Current Opinion in Biotechnology, 49,
148–155. https://doi.org/10.1016/j.copbio.2017.08.005
Gallone, B., Steensels, J., Prahl, T., Soriaga, L., Saels, V., Herrera‐Malaver, B.,
… Verstrepen, K. (2016). Domestication and divergence of Saccharomy-
ces cerevisiae beer yeasts. Cell, 166, 1397–1410.e16
CUBILLOS ET AL.
Garavaglia, C., & Swinnen, J. (2018). Economics of the craft beer revolu-
tion: A comparative international perspective. In C. Garavaglia, & J.
Swinner (Eds.), Economic perspectives on craft beer (pp. 3–51). Cham,
Switzerland: Springer Nature. https://doi.org/10.1007/978‐3‐319‐
58235‐1_1
Gayevskiy, V., & Goddard, M. R. (2016). Saccharomyces eubayanus and Sac-
charomyces arboricola reside in North Island native New Zealand
forests. Environmental Microbiology, 18, 1137–1147. https://doi.org/
10.1111/1462‐2920.13107
Gibson, B., Geertman, J.‐M. A., Hittinger, C. T., Krogerus, K., Libkind, D.,
Louis, E. J., … Sampaio, J. P. (2017). New yeasts—New brews: Modern
approaches to brewing yeast design and development. FEMS Yeast
Research, 17(4), fox038. https://doi.org/10.1093/femsyr/fox038
Gibson, B., Krogerus, K., Ekberg, J., Mikkelson, A., Pentikäinen, S., Wilpola,
A., and Vidgren, V. (2015). Non‐conventional yeast as a new tool for
beer flavour modification. Proceedings of the 35th European Brewing
Convention Congress, Porto, Portugal.
Gibson, B., & Liti, G. (2015). Saccharomyces pastorianus: Genomics insights
inspiring innovation for industry. Yeast, 32, 17–27. https://doi.org/
10.1002/yea.3033
Gibson, B. R., Storgårds, E., Krogerus, K., & Vidgren, V. (2013). Comparative
physiology and fermentation performance of Saaz and Frohberg lager
yeast strains and the parental species Saccharomyces eubayanus. Yeast,
30, 255–266. https://doi.org/10.1002/yea.2960
Glushakova, A. M., Ivannikova, Y. V., Naumova, E. S., Chernov, I. &
Naumov, G. I. (2007). Massive isolation and identification of Saccharo-
myces paradoxus yeasts from plant phyllosphere. Mikrobiologiya, 76(2),
236–242.
Gonçalves, M., Pontes, A., Almeida, P., Barbosa, R., Serra, M., Libkind, D., …
Sampaio, J. P. (2016). Distinct domestication trajectories in top‐
fermenting beer yeasts and wine yeasts. Current Biology, 26,
2750–2761. https://doi.org/10.1016/j.cub.2016.08.040
Gonzalez, S. S., Barrio, E., & Querol, A. (2008). Molecular characterization
of new natural hybrids of Saccharomyces cerevisiae and S. kudriavzevii
in brewing. Applied and Environmental Microbiology, 74, 2314–2320.
https://doi.org/10.1128/AEM.01867‐07
Gray, J. C., & Goddard, M. R. (2012). Sex enhances adaptation by unlinking
beneficial from detrimental mutations in experimental yeast popula-
tions. BMC Evolutionary Biology, 12, 43. https://doi.org/10.1186/
1471‐2148‐12‐43
Gschaedler, A. (2017). Contribution of non‐conventional yeasts in alcoholic
beverages. Current Opinion in Food Science, 13, 73–77. https://doi.org/
10.1016/j.cofs.2017.02.004
Gutiérrez, A., Boekhout, T., Gojkovic, Z., & Katz, M. (2018). Evaluation of
non‐Saccharomyces yeasts in the fermentation of wine, beer and cider
for the development of new beverages. Journal of the Institute of
Brewing, in press, 124, 389–402. https://doi.org/10.1002/jib.512
Haehn, H., & Glaubitz, M. (1933). Beer manufacture. Patent US1898047 A.
Hassan, Y. I., & Bullerman, L. B. (2008). Antifungal activity of Lactobacillus
paracasi ssp. tolerans isolated from a sourdough bread culture. Interna-
tional Journal of Food Microbiology, 121, 112–115. https://doi.org/
10.1016/j.ijfoodmicro.2007.11.038
Hebly, M., Brickwedde, A., Bolat, I., Driessen, M. R., de Hulster, E. A., van
den Broek, M., … Daran‐Lapujade, P. (2015). S. cerevisiae × S. eubayanus
interspecific hybrid, the best of both worlds and beyond. FEMS Yeast
Research, 15(3), fov005. fov005
Hittinger, C. T., Steele, J. L., & Ryder, D. S. (2018). Diverse yeasts for diverse
fermented beverages and foods. Current Opinion in Biotechnology, 49,
199–206. https://doi.org/10.1016/j.copbio.2017.10.004
Holt, S., Mukherjee, V., Lievens, B., Verstrepen, K. J., & Thevelein, J. M.
(2018). Bioflavoring by non‐conventional yeasts in sequential beer
395
fermentations. Food Microbiology, 72, 55–66. https://doi.org/
10.1016/j.fm.2017.11.008
Hranilovic, A., Gambetta, J. M., Schmidtke, L., Boss, P. K., Grbin, P. R.,
Masneuf‐Pomarede, I., … Jiranek, V. (2018). Oenological traits of
Lachancea thermotolerans show signs of domestication and allopatric
differentiation. Nature Scientific Reports, 8, 14812. https://doi.org/
10.1038/s41598‐018‐33105‐7
Huige, N. J., Sanchez, G. W., & Leidig, A. R. (1990). U.S. Patent No.
4,970,082. Washington, DC: U.S. Patent and Trademark Office.
Jespersen, L. (2003). Occurrence and taxonomic characteristics of strains
of Saccharomyces cerevisiae predominant in African indigenous
fermented foods and beverages. FEMS Yeast Research, 3(2), 191–200.
https://doi.org/10.1016/S1567‐1356(02)00185‐X
Kapsopoulou, K., Kapaklis, A., & Spyropoulos, H. (2005). Growth and fer-
mentation characteristics of a strain of the wine yeast Kluyveromyces
thermotolerans isolated in Greece. World Journal of Microbiology and
Biotechnology, 21, 1599–1602. https://doi.org/10.1007/s11274‐005‐
8220‐3
Krogerus, K., Holmström, S., & Gibson, B. (2018). Enhanced wort fermenta-
tion with de novo lager hybrids adapted to high‐ethanol environments.
Applied and Environmental Microbiology, 84(4), 1–20.
Krogerus, K., Magalhães, F., Vidgren, V., & Gibson, B. (2015). New lager
yeast strains generated by interspecific hybridization. Journal of Indus-
trial Microbiology and Biotechnology, 42(5), 769–778. https://doi.org/
10.1007/s10295‐015‐1597‐6
Krogerus, K., Magalhães, F., Vidgren, V., & Gibson, B. (2017). Novel
brewing yeast hybrids: Creation and application. Applied Microbiology
and Biotechnology, 101(1), 65–78. https://doi.org/10.1007/s00253‐
016‐8007‐5
Krogerus, K., Preiss, R., & Gibson, B. (2018). A unique Saccharomyces
cerevisiae × Saccharomyces uvarum hybrid isolated from Norwegian
farmhouse beer: Characterization and reconstruction. Frontiers in
Microbiology, in press, 9, 2253.
Krogerus, K., Seppanen‐Laakso, T., Castillo, S., & Gibson, B. (2017). Inheri-
tance of brewing‐relevant phenotypes in constructed Saccharomyces
cerevisiae × Saccharomyces eubayanus hybrids. Microbial Cell Factories,
16, 66. https://doi.org/10.1186/s12934‐017‐0679‐8
Laitila, A., Wilhelmson, A., Kotaviita, E., Olkku, J., Home, S., & Juvonen, R.
(2006). Yeasts in an industrial malting ecosystem. Journal of Industrial
Microbiology and Biotechnology, 33(11), 953–966. https://doi.org/
10.1007/s10295‐006‐0150‐z
Lappe‐Oliveras, P., Moreno‐Terrazas, R., Arrizón‐Gaviño, J., Herrera‐
Suárez, T., García‐Mendoza, A., & Gschaedler‐Mathis, A. (2008). Yeasts
associated with the production of Mexican alcoholic nondistilled and
distilled Agave beverages. FEMS Yeast Research, 8(7), 1037–1052.
https://doi.org/10.1111/j.1567‐1364.2008.00430.x
Legras, J.‐L., Galeote, V., Bigey, F., Camarasa, C., Marsit, S., Nidelet, T., …
Dequin, S. (2018). Adaptation of S. cerevisiae to fermented food envi-
ronments reveals remarkable genome plasticity and the footprints of
domestication. Molecular Biology and Evolution, 35, 1712–1727.
https://doi.org/10.1093/molbev/msy066
Legras, J.‐L., Merdinoglu, D., Cornuet, J.‐M., & Karst, F. (2007). Bread, beer
and wine: Saccharomyces cerevisiae diversity reflects human history.
Molecular Ecology, 16, 2091–2102. https://doi.org/10.1111/j.1365‐
294X.2007.03266.x
Li, H., Liu, Y., & Zhang, W. (2011). Method for preparing non‐alcoholic beer
by Candida shehatae. Patent CN102220198 B.
Libkind, D., Hittinger, C. T., Valerio, E., Gonçalves, C., Dover, J., Johnston,
M., … Sampaio, J. P. (2011). Microbe domestication and the identifi-
cation of the wild genetic stock of lager‐brewing yeast. Proceedings
of the National Academy of Sciences of the United States of
396
America, 108(35), 14539–14544. https://doi.org/10.1073/pnas.110
5430108
Loponen, J., & Sibakov, J. (2013). Sourdough and cereal beverages. In M.
Gobbetti, & M. Gänzle (Eds.), Handbook on sourdough biotechnology.
New York, USA: Springer. https://doi.org/10.1007/978‐1‐4614‐
5425‐0_11
Lutz, H. F. (1922). In J. C. Hinrich, & G. E. Stechert (Eds.), Viticulture and
brewing in the ancient Orient. Leipzig, Germany: Applewood Books.
https://doi.org/10.5962/bhl.title.23430
Lyumugabe, F., Gros, J., Nzungize, J., Bajyana, E., & Thonart, P. (2012).
Characteristics of African traditional beers brewed with sorghum malt:
A review. Biotechnology, Agronomy, Society and Environment, 16(4),
509–530.
Marongiu, A., Zara, G., Legras, J.‐L., Del Caro, A., Mascia, I., Fadda, C., &
Budroni, M. (2015). Novel starters for old processes: Use of Saccharomy-
ces cerevisiae strains isolated from artisanal sourdough for craft beer
production at a brewery scale. Journal of Industrial Microbiology and Bio-
technology, 42, 85–92. https://doi.org/10.1007/s10295‐014‐1525‐1
Marshall, E., & Mejia, D. (2012). Traditional fermented food and beverages
for improved livelihoods. FAO Diversification booklet 21.
Mascia, I., Fadda, C., Dostálek, P., Kerabín, M., Zara, G., Budroni, M., & Del
Caro, A. (2015). Is it possible to create an innovative craft durum wheat
beer with sourdough yeasts? A case study. Journal of the Institute of
Brewing, 121, 283–286. https://doi.org/10.1002/jib.215
Mascia, I., Fadda, C., Kerabín, M., Dostálek, P., & Del Caro, A. (2016). Aging
of craft durum wheat beer fermented with sourdough yeasts. LWT ‐
Food Science and Technology, 65, 487–494. https://doi.org/10.1016/j.
lwt.2015.08.026
Mendoza, L. M., Neef, A., Vignolo, G., & Belloch, C. (2017). Yeast diversity
during the fermentation of Andean chicha: A comparison of high‐
throughput sequencing and culture‐dependent approaches. Food
Microbiology, 67, 1–10. https://doi.org/10.1016/j.fm.2017.05.007
Mertens, S., Steensels, J., Saels, V., De Rouck, G., Aerts, G., & Verstrepen,
K. J. (2015). A large set of newly created interspecific Saccharomyces
hybrids increases aromatic diversity in lager beers. Applied and Environ-
mental Microbiology, 81, 8202–8214. https://doi.org/10.1128/
AEM.02464‐15
Michel, M., Kopecká, J., Meier‐Dörnberg, T., Zarnkow, M., Jacob, F., &
Hutzler, M. (2016). Screening for new brewing yeasts in the
non‐Saccharomyces sector with Torulaspora delbrueckii as model. Yeast,
33, 129–144. https://doi.org/10.1002/yea.3146
Mora, J., Barbas, J. I., & Mulet, A. (1990). Growth of yeast species during
the fermentation of musts inoculated with Kluyveromyces
thermotolerans and Saccharomyces cerevisiae. American Journal of Enol-
ogy and Viticulture, 41, 156–159.
Moroni, A. V., Arendt, E. K., & Dal Bello, F. (2011). Biodiversity of lactic
acid bacteria and yeasts in spontaneously‐fermented buckwheat and
teff sourdoughs. Food Microbiology, 28, 497–502.
Mukai, N., Masaki, K., Fujii, T., Kawamukai, M., & Iefuji, H. (2010). PAD1
and FDC1 are essential for the decarboxylation of phenylacrylic acids
in Saccharomyces cerevisiae. Journal of Bioscience and Bioengineering,
109, 564–569. https://doi.org/10.1016/j.jbiosc.2009.11.011
Mukherjee, V., Radecka, D., Aerts, G., Verstrepen, K. J., Lievens, B., &
Thevelein, J. M. (2017). Phenotypic landscape of non‐conventional
yeast species for different stress tolerance traits desirable in bioethanol
fermentation. Biotechnology for Biofuels, 10(1), 216. https://doi.org/
10.1186/s13068‐017‐0899‐5
Naseeb, S., Alsammar, H., Burgis, T., Donaldson, I., Knyazev, N., Knight, C.,
& Delneri, D. (2018). Whole genome sequencing, de novo assembly
and phenotypic profiling for the new budding yeast species Saccharo-
myces jurei. G3: Genes, Genomes, Genetics, 8, 2967–2977.
CUBILLOS ET AL.
N'Guessan, F. K., N'Dri, Y. N., Camara, F., & Djè, M. K. (2010). Saccharomy-
ces cerevisiae and Candida tropicalis as starter cultures for the alcoholic
fermentation of tchapalo, a traditional sorghum beer. World Journal of
Microbiology and Biotechnology, 26, 693–699. https://doi.org/
10.1007/s11274‐009‐0224‐y
Nguyen, H. V., & Boekhout, T. (2017). Characterization of Saccharomyces
uvarum (Beijerinck, 1898) and related hybrids: Assessment of molecular
markers that predict the parent and hybrid genomes and a proposal to
name yeast hybrids. FEMS Yeast Research, 17(2), fox014.
Nikulin, J., Krogerus, K., & Gibson, B. (2018). Alternative Saccharomyces
interspecies hybrid combinations and their potential for low‐
temperature wort fermentation. Yeast, 35, 113–127. https://doi.org/
10.1002/yea.3246
Nyanga, L. K., Nout, M. J. R., Gadaga, T. H., Theelen, B., Boekhout, T., &
Zwietering, M. H. (2007). Yeasts and lactic acid bacteria microbiota
from masau (Ziziphus mauritania) fruits and their fermented fruit pulp
in Zimbabwe. International Journal of Food Microbiology, 120,
159–166. https://doi.org/10.1016/j.ijfoodmicro.2007.06.021
Osburn, K., Ahmad, N. N., & Bochman, M. L. (2016). Bio‐prospecting, selection,
and analysis of wild yeasts for ethanol fermentation. Zymurgy, 39, 81–88.
Osburn, K., Amaral, J., Metcalf, S. R., Nickens, D. M., Rogers, C. M., Sausen,
C., … Bochman, M. L. (2018). Primary souring: A novel bacteria‐free
method for sour beer production. Food Microbiology, 70, 76–84.
https://doi.org/10.1016/j.fm.2017.09.007
Palomba, S., Blaiotta, G., Ventorino, V., Saccone, A., & Pepe, O. (2011).
Microbial characterization of sourdough for sweet baked products in
the Campania region (southern Italy) by a polyphasic approach. Annals
of Microbiology, 61, 307–314.
Paredes, R. S., Vieira, I. V., de Mello, V., Vilela, L., Schwan, R., & Eleutherio,
E. (2018). Identification of three robust and efficient Saccharomyces
cerevisiae strains isolated from Brazilian's cachaça distilleries. Biotech-
nology Research and Innovation, in press, 2(1), 22–29.
Peris, D., Belloch, C., Lopandic, K., Alvarez‐Perez, J. M., Querol, A., & Bar-
rio, E. (2012). The molecular characterization of new types of
Saccharomyces cerevisiae × S. kudriavzevii hybrid yeasts unveils a high
genetic diversity. Yeast, 29, 81–91. https://doi.org/10.1002/yea.2891
Peris, D., Langdon, Q. K., Moriarty, R. V., Sylvester, K., Bontrager, M.,
Charron, G., … Hittinger, C. T. (2016). Complex ancestries of lager‐
brewing hybrids were shaped by standing variation in the wild yeast
Saccharomyces eubayanus. PLoS Genetics, 12, e1006155. https://doi.
org/10.1371/journal.pgen.1006155
Peris, D., Sylvester, K., Libkind, D., Gonçalves, P., Sampaio, J. P., Alexander,
W. G., & Hittinger, C. T. (2014). Population structure and reticulate evo-
lution of Saccharomyces eubayanus and its lager‐brewing hybrids.
Molecular Ecology, 23, 2031–2045. https://doi.org/10.1111/mec.12702
Peter, J., De Chiara, M., Friedrich, A., Yue, J.‐X., Pflieger, D., Bergström, A.,
… Cruaud, C. (2018). Genome evolution across 1,011 Saccharomyces
cerevisiae isolates. Nature, 556, 339–344. https://doi.org/10.1038/
s41586‐018‐0030‐5
Piló, F., Carvajal‐Barriga, E., Guamán‐Burneo, M., Portero‐Barahona, P.,
Dias, A., & de Freitas, L. Gomes F., & Rosa, C. (2018). Saccharomyces
cerevisiae populations and other yeasts associated with indigenous
beers (chicha) of Ecuador. Brazilian Journal of Microbiology, in press,
49, 808–815. https://doi.org/10.1016/j.bjm.2018.01.002
Preiss, R., Tyrawa, C., Krogerus, K., Garshol, L. M., & Van Der Merwe, G.
(2018). Traditional Norwegian kveik are a genetically distinct group of
domesticated Saccharomyces cerevisiae brewing yeasts. Frontiers in
Microbiology, 9, 2137. https://doi.org/10.3389/fmicb.2018.02137
Pulvirenti, A., Solieri, L., Gullo, M., De Vero, L., & Giudici, P. (2004). Occurrence
and dominance of yeast species in sourdough. Letters in Applied Microbiol-
ogy, 38, 113–117. https://doi.org/10.1111/j.1472‐765X.2003.01454.x
CUBILLOS ET AL.
Ramos, C. L., & Schwan, R. F. (2017). Technological and nutritional aspects
of indigenous Latin America fermented foods. Current Opinion in Food
Science, 13, 97–102. https://doi.org/10.1016/j.cofs.2017.07.001
Ravasio, D., Carlin, S., Boekhout, T., Groenewald, M., Vrhovsek, U.,
Walther, A., & Wendland, J. (2018). Adding flavor to beverages with
non‐conventional yeasts. Fermentation, 4(1), 15. https://doi.org/
10.3390/fermentation4010015
Richard, P., Viljanen, K., & Penttila, M. (2015). Overexpression of PAD1 and
FDC1 results in significant cinnamic acid decarboxylase activity in Sac-
charomyces cerevisiae. AMB Express, 5, 12. https://doi.org/10.1186/
s13568‐015‐0103‐x
Ripari, V., Tomassetti, M., Cecchi, T., & Enrico, B. (2018). Recipe, volatiles
profile, sensory analysis, physico‐chemical and microbial characteriza-
tion of acidic beers from both sourdough yeasts and lactic acid
bacteria. European Food Research and Technology, 244(11), 2027–2040.
Rodriguez, M. E., Origone, A. C., Flores, M. G., & Lopez, C. A. (2016). Sac-
charomyces in traditional and indigenous fermentations from
Patagonia. In N. Olivera, D. Libkind, & E. Donati (Eds.), Biology and bio-
technology of Patagonian microorganisms (pp. 251–276). Cham:
Springer. https://doi.org/10.1007/978‐3‐319‐42801‐7_15
Rodriguez, M. E., Perez‐Traves, L., Sangorrin, M. P., Barrio, E., & Lopes, C.
A. (2014). Saccharomyces eubayanus and Saccharomyces uvarum associ-
ated with the fermentation of Araucaria araucana seeds in Patagonia.
FEMS Yeast Research, 14, 948–965. https://doi.org/10.1111/1567‐
1364.12183
Rossi, S., Turchetti, B., Sileoni, V., Marconi, O., & Perretti, G. (2018). Evalu-
ation of Saccharomyces cerevisiae strains isolated from non‐brewing
environments in beer production. Journal of the Institute of Brewing.,
124, 381–388. https://doi.org/10.1002/jib.503
Rosso, L., Lobry, J., Bajard, S., & Flandrois, J. (1995). Convenient model to
describe the combined effects of temperature and pH on microbial
growth. Applied and Environmental Microbiology, 61, 610–616.
Russell, J., & Diez‐Gonzalez, F. (1997). The effects of fermentation acids on
bacterial growth. Advances in Microbial Physiology, 39, 205–234.
https://doi.org/10.1016/S0065‐2911(08)60017‐X
Ruyters, S., Mukherjee, V., Verstrepen, K. J., Thevelein, J. M., Willems, K.
A., & Lievens, B. (2015). Assessing the potential of wild yeasts for
bioethanol production. Journal of Industrial Microbiology & Biotechnol-
ogy, 42(1), 39–48. https://doi.org/10.1007/s10295‐014‐1544‐y
Saerens, S., & Swiegers, J. H. (2017). U.S. Patent Application No.
15/413,246.
Sampaio, J. P., & Gonçalves, P. (2008). Natural populations of Saccharomyces
kudriavzevii in Portugal are associated with oak bark and are sympatric
with S. cerevisiae and S. paradoxus. Applied and Environmental Microbiol-
ogy, 74(7), 2144–2152. https://doi.org/10.1128/AEM.02396‐07
Schoch, C., Seifert, K., Huhndorf, S., Robert, V., Spouge, J., Levesque, C., …
Fungal Barcoding Consortium (2012). Nuclear ribosomal internal tran-
scribed spacer (ITS) region as a universal DNA barcode marker for
Fungi. Proceedings of the National Academy of Sciences, 109(16),
6241–6246.
Segura‐García, L. E., Taillandier, P., Brandam, C., & Gschaedler, A. (2015).
Fermentative capacity of Saccharomyces and non‐Saccharomyces in
Agave juice and semi‐synthetic medium. LWT ‐ Food Science and Tech-
nology, 60(1), 284–291. https://doi.org/10.1016/j.lwt.2014.08.005
Senkarcinova, B., Dias, I. A. G., Nespor, J., & Branyik, T. (2019). Probiotic
alcohol‐free beer made with Saccharomyces cerevisiae var. boulardii.
LWT ‐ Food Science and Technology, 100, 362–367. https://doi.org/
10.1016/j.lwt.2018.10.082
Sipiczki, M. (2016). Overwintering of vineyard yeasts: Survival of
interacting yeast communities in grapes mummified on vines. Frontiers
in Microbiology, 7, 212.
397
Sniegowski, P. D., Dombrowski, P. G., & Fingerman, E. (2002). Saccharomy-
ces cerevisiae and Saccharomyces paradoxus coexist in a natural
woodland site in North America and display different levels of repro-
ductive isolation from European conspecifics. FEMS Yeast Research, 1,
299–306. https://doi.org/10.1111/j.1567‐1364.2002.tb00048.x
Spitaels, F., Wieme, A. D., Janssens, M., Aerts, M., Daniel, H. M., Van
Landschoot, A., … Vandamme, P. (2014). The microbial diversity of tra-
ditional spontaneously fermented lambic beer. PLoS One, 9(4), e95384.
https://doi.org/10.1371/journal.pone.0095384
Spitaels, F., Wieme, A. D., Janssens, M., Aerts, M., Van Landschoot, A., De
Vuyst, L., & Vandamme, P. (2015). The microbial diversity of an indus-
trially produced lambic beer shares members of a traditionally
produced one and reveals a core microbiota for lambic beer fermenta-
tion. Food Microbiology, 49, 23–32. https://doi.org/10.1016/j.
fm.2015.01.008
Steensels, J., Meersman, E., Snoek, T., Saels, V., & Verstrepen, K. J. (2014).
Large‐scale selection and breeding to generate industrial yeasts with
superior aroma production. Applied and Environmental Microbiology,
80, 6965–6975. https://doi.org/10.1128/AEM.02235‐14
Steensels, J., Snoek, T., Meersman, E., Picca Nicolino, M., Voordeckers, K.,
& Verstrepen, K. J. (2014). Improving industrial yeast strains: Exploiting
natural and artificial diversity. FEMS Microbiology Reviews, 38,
947–995. https://doi.org/10.1111/1574‐6976.12073
Takao, Y., Takahashi, T., Yamada, T., Goshima, T., Isogai, A., Sueno, K., …
Akao, T. (2018). Characteristic features of the unique house sake yeast
strain Saccharomyces cerevisiae Km67 used for industrial sake brewing.
Journal of Bioscience and Bioengineering, in press, 126, 617–623.
https://doi.org/10.1016/j.jbiosc.2018.05.008
Trindade de Carvalho, B., Holt, S., Souffriau, B., Lopes Brandao, R.,
Foulquie‐Moreno, M. R., & Thevelein, J. M. (2017). Identification of
novel alleles conferring superior production of rose flavor phenylethyl
acetate using polygenic analysis in yeast. MBio, 8. https://doi.org/
10.1128/mBio.01173‐17
Vallejo, J. A., Miranda, P., Flores‐Félix, J. D., Sánchez‐Juanes, F., Ageitos, J.
M., González‐Buitrago, J. M., … Villa, T. G. (2013). Atypical yeasts iden-
tified as Saccharomyces cerevisiae by MALDI‐TOF MS and gene
sequencing are the main responsible of fermentation of chicha, a tradi-
tional beverage from Peru. Systematic and Applied Microbiology, 36(8),
560–564. https://doi.org/10.1016/j.syapm.2013.09.002
van Rijswijck, I. M. H., Wolkers‐Rooijackers, J. C. M., Abee, T., & Smid, E. J.
(2017). Performance of non‐conventionoal yeasts in co‐culture with
brewer's yeasts for steering ethanol and aroma production. Microbial
Biotechnology, 10, 1591–1602.
Varela, C. (2016). The impact of non‐Saccharomyces yeasts in the produc-
tion of alcoholic beverages. Applied Microbiology and Biotechnology,
100(23), 9861–9874. https://doi.org/10.1007/s00253‐016‐7941‐6
Vidgren, V., & Londesborough, J. (2011). 125th anniversary review: Yeast
flocculation and sedimentation in brewing. Journal of the Institute of
Brewing, 117, 475–487. https://doi.org/10.1002/j.2050‐0416.2011.
tb00495.x
Viigand, K., Põšnograveja, K., Visnapuu, T., & Alamäe, T. (2018). Genome
mining of non‐conventional yeasts: Search and analysis of MAL clus-
ters and proteins. Genes, 9, 354. https://doi.org/10.3390/
genes9070354
Vogelmann, S. A., & Hertel, C. (2011). Impact of ecological factors on the
stability of microbial associations in sourdough fermentation. Food
Microbiology, 28, 583–589. https://doi.org/10.1016/j.fm.2010.11.010
Vrancken, G., De Vuyst, L., Van der Meulen, R., Huys, G., Vandamme, P., &
Daniel, H. M. (2010). Yeast species composition differs between artisan
bakery and spontaneous laboratory sourdoughs. FEMS Yeast Research,
10, 471–481. https://doi.org/10.1111/j.1567‐1364.2010.00621.x
398
Wang, Q. M., Liu, W. Q., Liti, G., Wang, S. A., & Bai, F. Y. (2012). Surpris-
ingly diverged populations of Saccharomyces cerevisiae in natural
environments remote from human activity. Molecular Ecology, 21(2),
5404–5417. https://doi.org/10.1111/j.1365‐294X.2012.05732.x
Wang, S. A., & Bai, F. Y. (2008). Saccharomyces arboricolus sp. nov., a yeast
species from tree bark. International Journal of Systematic and Evolution-
ary Microbiology, 58, 510–514. https://doi.org/10.1099/ijs.0.65331‐0
Warringer, J., Zorgo, E., Cubillos, F. A., Zia, A., Gjuvsland, A., Simpson, J. T.,
… Blomberg, A. (2011). Trait variation in yeast is defined by population
history. PLoS Genetics, 7(6), e1002111. https://doi.org/10.1371/jour-
nal.pgen.1002111
White, C., & Zainasheff, J. (2010). Yeast: The practical guide to beer fermen-
tation. Boulder, CO, USA: Brewers Publications.
CUBILLOS ET AL.
Zhang, H., Skelton, A., Gardner, R. C., & Goddard, M. R. (2010). Saccharo-
myces paradoxus and Saccharomyces cerevisiae reside on oak trees in
New Zealand: Evidence for migration from Europe and interspecies
hybrids. FEMS Yeast Research, 10, 941–947. https://doi.org/10.1111/
j.1567‐1364.2010.00681.x
How to cite this article: Cubillos FA, Gibson B, Grijalva‐
Vallejos N, Krogerus K, Nikulin J. Bioprospecting for brewers:
Exploiting natural diversity for naturally diverse beers. Yeast.
2019;36:383–398. https://doi.org/10.1002/yea.3380