Screening for productive strains and

strain improvement in biotechnological

organisms

Chapter 2
 Introduction
 All living things are composed of cells, of which there are two basic
types, the prokaryotic cell and the eucaryotic cell.
 Cell wall: Procaryotic cell walls contain glycopeptides; these are
absent in eucaryotic cells.
 Cell walls of eucaryotic cells contain chitin, cellulose and other
sugar polymers.
These provide rigidity where cell walls are present.
 What kinds of microorganism ?
• Bacteria
• Fungi (Yeast and molds)
• Viruses
• Protozoans, algae, Helminths (worms) to lesser
extent
• Protozoans and helminths are considered
Accidental”
Classification of organisms
•
 Cont’
• Plantae (multicellular, eukaryotic)
• Animalia (multicellular, eukaryotic)
• Fungi (multicellular, eukaryotic)
• Protista ( Eukaryotic, unicellular and

multicellular)
• Eubacteria (Unicellular, prokaryotic)
• Archaebacteria (Unicellular, prokaryotic )

Where are viruses and prions? A prion is a type of

protein that can cause disease in animals and
humans by triggering normally healthy proteins in
the brain to fold abnormally.
 The 3 domains

1. Eukarya
Have a nucleus & organelles (humans, animals,
plants, fungi and protists)

2. Eubacteria
True bacteria, peptidoglycan

3. Archaea
• With unusual cell walls, and membreanes
• odd bacteria that live in extreme environments,
high salt, heat, etc. (usually called
extremophiles)
 Cell type
Prokaryotes Eukaryotes
Larger cells
• Smaller cells -Cell have nucleus &
• No nucleus or organelles
organelles
• Single –celled
 Can be single celled or
Bacteria and archaea
multi
 Plantae, Fungi
• Viruses and prions are not Animalia,
cells so are not considered and , Protista
alive.
 IMPORTANT IN INDUSTRIAL MICROBIOLOGY AND
BIOTECHNOLOGY
• Microorganisms have the following
advantages over plants or animals as inputs in
biotechnology:
 MOS grow rapidly in comparison with plants and
animals. The generation time (the time for an
organism to mature and reproduce) is about
12 years in man, about 24 months in cattle, 18
months in pigs, 6 months in chicken,
but only 15 minutes in the bacterium, E coli. The
consequence is that biotechnological products
which can be obtained from MOS in a matter of
days may take many months in animals or plants.
 IMPORTANT IN INDUSTRIAL MICROBIOLOGY AND
BIOTECHNOLOGY
 The space requirement for growth MOs is small.
 MOS are not subject to the problems of the

vicissitudes of weather which may affect

agricultural production especially among plants.
 MOs are not affected by diseases of plants and

animals, although they do have their peculiar

scourges in the form phages and contaminants,
but there are procedure to contain them.
 Source of MOs used in Biotechnology

 Although the well-known ubiquity of MO implies

that almost any natural ecological entity–water,
air, leaves, tree trunks – may provide MOs, the
soil is the preferred source for isolating organisms,
because it is a vast reservoir of diverse organisms.
 Indeed MOs capable of utilizing virtually any

carbon source will be found in soil if adequate

screening methods are used.
 In recent times, other ‘new’ habitats, especially the

marine environment, have been included in

habitats to be studied in searches for bioactive
microbial metabolites or ‘bio-mining’.
ISOLATION, SCREENING AND STRAIN
IMPROVEMENT
 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

 The diversity of MOs may exploited still by

searching for strains from the natural
environment able to produce products of
commercial value.
 The first stage in the screening for microorganism

of potential industrial application is their isolation

 Isolation involves obtaining either pure or mixed

cultures followed by their assessment to
determine which carry out the desired reaction
or produce the desired product.
 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

Culture Dependent Microbial Community Analysis

Isolation :-The separation of individual
organisms from the mixed community
Enrichment Cultures :-Select for desired
organisms through manipulation of
medium and incubation conditions
Inocula :-The sample from which
microorganisms will be isolated
 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

Criteria as being important in the choice of organism:

1. The nutritional characteristics of the organism. It is
frequently required that a process be carried out using
a very cheap medium or a pre-determined one, e.g. the
use of methanol as an energy source. These
requirements may be met by the suitable design of the
isolation medium.
2. The optimum temperature of the organism. The use of
an organism having an optimum temperature above
40° considerably reduces the cooling costs of a large-
scale fermentation and, therefore, the use of such a
temperature in the isolation procedure may be
beneficial.
 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

Criteria as being important in the choice of organism:

3. The reaction of the organism with the equipment to be
employed and the suitability of the organism to the type of
process to be used.
4. The stability of the organism and its amenability to
genetic manipulation.
5. The productivity of the organism, measured in its ability
to convert substrate into product and to give a high yield
of product per unit time.
6. The ease of product recovery from the culture
 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

Isolation and Screening of Industrial Strain

Isolation from the environment is by:-
 Collecting samples of free living microorganism from

anthropogenic or natural habitats.

 These isolates are then screened for desirable traits.

 Or by sampling from specific sites:

 Most with desired characteristics are found among the

natural microflora
 After sampling of the organism the next step is of

enrichment.
 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

Conventional bacteriological-based methods

 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

Isolation and Screening of Industrial Strain

Screening
 Subsequent isolation as pure cultures on solid

growth media involves choosing or developing the

appropriate selective media and growth conditions.
 Next step to enrichment and isolation is

Screening.
 The pure cultures must be screened for the desired

property; production of a specific enzyme,

inhibitory compound, etc.
 Selected isolates must also be screened for other

important features, such as stability and, where

necessary, non-toxicity.
 ISOLATION, SCREENING AND STRAIN IMPROVEMENT

Screening
These isolation and screening procedures are more easily,
applied to the search for a single microorganism.
The industrial microorganism should ideally exhibit:
1. Genetic stability

2. Efficient production of the target product, whose, route

of biosynthesis, should preferably be well characterized.
3. Limited or no need for vitamins and additional growth
factors.
4. Utilization of a wide range of low-cost and readily
available carbon sources
5. Amenability to genetic manipulation;
6. Safety, non-pathogenic and should not produce toxic
agents, unless there is the target product;
7. Ready harvesting from the fermentation;
 ROUTINE LABORATORY MEDIA

These are classified into six types:

(1) Basal media, (2) Enriched media,
(3) Selective media (4) Indicator media,
(5) Transport media, and (6) Storage media.
(1) BASAL MEDIA, basically simple media that supports most non-fastidious
bacteria.
Examples: Nutrient broth, nutrient agar and peptone water. Staphylococcus
and Enterobacteriaceae grow in these media.
2. ENRICHED MEDIA, are used to grow nutritionally exacting (fastidious)
bacteria . The basal media are enriched usually by adding blood, serum or
egg.
Examples: blood agar , chocolate agar, Loeffler’s serum slope and
Lowenstein-Jensen media. Streptococci grow in blood agar media.
(3) SELECTIVE MEDIA. Favour the growth of a particular bacterium by
inhibiting the growth of undesired bacteria and allowing growth of desirable
bacteria.
Examples: MacConkey agar, Lowenstein-Jensen media, tellurite media
Antibiotic may be added to a medium for inhibition.
 Cont…Selective
 Some ingredient (such as an unusual sugar) that favors
the growth of the targeted organism is included in the
medium.
 The medium might also contain ingredients (such as
antibiotics, salt, surfactants, etc.) that inhibit the
growth of competing bacteria.
 Table 2.1 gives some examples of selective media.
 Cont….Selective Media
 In the case of Baird- Parker agar (see Fig. below), lithium
chloride, glycine, and tellurite allow S. aureus to grow
while suppressing a larger number of other organisms.

 However, if injured , target organisms may be killed by

the selective agent.
 In addition, the selective media are not entirely selective.

 Lactobacillus de Man-Rogosa-Sharpe (MRS) medium is

selective for lactobacilli but also supports the growth of

Listeria monocytogenes.
 ROUTINE LABORATORY MEDIA

(4). INDICATOR (DIFFERENTIAL) MEDIA, distinguish one

microorganism type from another growing on the same media.
 This type of media uses the biochemical characteristics of a

microorganism growing in the presence of specific nutrients

or indicators (such as neutral red, phenol red or methylene
blue) added to the medium to visibly indicate the defining
characteristics of a microorganism.
 When a particular substrate (carbohydrate) is incorporated

into a medium and a mixture of bacteria inoculated on it,

only that bacterium that can ferment it produces acid.
 This change in pH is detected by using a pH indicator

incorporated in the medium and the bacterium that can

ferment the sugar appears in a different colour. This
approach is used in MacConkey’s agar, CLED agar, TCBS
agar, XLD agar etc.
 ROUTINE LABORATORY MEDIA

 MacConkey’s agar is the most commonly used media to

culture and identify gram negative bacilli (especially
enterobacteriaceae members).
 It contains bile salts (selective agent), lactose (sugar),
peptone and neutral red (pH indicator), agar and water.
 Those bacteria that can ferment lactose produce pink
coloured colonies where non-lactose fermenting colonies
produce colourless colonies.
 ROUTINE LABORATORY MEDIA
(5)TRANSPORT MEDIA, Clinical specimens must be
transported to the laboratory immediately after collection
to prevent overgrowth of contaminating organisms or
commensals.
 This can be achieved by using transport media. Such

media prevent drying (desiccation) of specimen,

maintain the viability of all organisms in the specimen
without altering their concentration .
Examples: Cary-Blair medium, Amies medium, Stuart
medium.

(6) STORAGE MEDIA. Media used for storing the bacteria

for a long period of time. Examples: Egg saline medium,
chalk cooked meat broth
 Isolation methods
Methods of isolating pure culture
 A culture contains more than one species of

microbes is a mixed culture.

 A culture containing only one species of

microbes is a pure culture.

The following methods are used to isolate pure
culture
1. Streak plate technique
2. Pour plate technique
3. Spread plate technique
 Isolation methods

1. Streak-plate method
 The process of spreading the microbial culture

with an inoculating needle on the surface of the

media.
 Sterilize the inoculating needle by flame to

make red hot and allow it to cool for 30 sec.

 Isolation methods

Streak-plate method
 Isolation methods

2. Pour-plate method
 The bacterial culture and liquid agar medium

are mixed together.

 After mixing the medium, the medium

containing the culture poured into sterilized

petridishes (petriplates), allowed solidifying and
then incubated.
 After incubation colonies appear on the surface.
 Isolation methods
 Pour-plate method
 Isolation methods
Disadvantage of Pour-plate method
1. The Mos are trapped beneath the surface of
medium when it solidifies. Hence, surface as
well as subsurface colonies are developed and it
is very difficult to isolate and count the surface
colonies.
2. This method is tedious, time consuming and
requires skill
3. The Mos are subjected to hot shock because
liquid medium is maintained at 45oC
temperature
4. This method is unsuitable for isolation of
psychrophile bacteria.
 Isolation methods
3. Spread plate technique
 this is the best method to isolate the pure

colonies.
 In this technique, the culture is not mixed with

the agar medium. Instead it is mixed with normal

saline and serially diluted.
 0.1 ml of sample taken from diluted mixture,

which is placed on the surface of agar plate and

spread evenly over the surface by using L shaped
glass rod called spreader.
 Incubate the plates
 After incubation, colonies are observed on the

agar surface.
 Isolation methods
3. Spread plate technique
 Isolation methods
Advantage of Spread plate method
1. It is a simple method
2. In this method only surface colonies are formed
3. Mos are not exposed to higher temperature
Pour plate Vs Spread plate
Enrichment and Isolation
 Enrichment culture is a technique resulting in
an increase in the number of a given organism
relative to the numbers of other types in the
original inoculum.
 Enrichment Cultures

• Can prove the presence of an organism in a

habitat
• Cannot prove an organism does not inhabit an
environment
 Enrichment and Isolation
Enrichment
 Enrichment in batch or continuous system on a

defined growth media and cultivation conditions

are performed to encourage the growth of the
organism with desired trait.
 This will increase the quantity of the desired

organism prior to isolation and screening.

Ex : Tetrathionate broth used for salmonella
typhi
Culture Preservation
 Culture Preservation
 Preservation of cultures by freezing, drying, or a combination
of the two processes is highly influenced by resistance of the
culture to the damage caused by rapid freezing, the
dehydrating effects of slow freezing, or damage caused during
recovery.
 To minimize damage, agents have been used that protect

against ice formation by causing the formation of glasses

upon cooling.
 Methods to protect against the negative effects of

dehydration include adaptation to lower effective water

activity by pre-incubation in high osmotic pressure solutions.
 Damage caused by thawing after freezing can be minimized

by rapid melting and by the composition of the medium used

for growth after preservation.
 Culture Preservation
 There are various preservation methods .
 Storage at reduced temperature agar, liquid
nitrogen
 Storage in dehydrated form dried culture,
lyophilization
 To date, preservation in liquid nitrogen is still the

most successful long-term method.

 Culture Preservation
Preservation in Distilled Water
 This method (Castellani method, 50 years ago) was

extensively tested on 594 fungal strains:

 62% of the strains growing and maintaining their

original morphology.
 In another study, 76% of yeasts, filamentous fungi, and

actinomycetes survived storage in distilled water for 10

years.
 The pathogen Sporothrix schencki concluded that even

though long-term survival was good when this procedure

was used, there was a noted loss in virulence.
 Castellani technique should be considered as one of the

options for practical storage of fungal isolates.

 Culture Preservation
Preservation under Oil
 One of the earlier preservation methods was the

use of mineral oil to prolong the utility of stock

cultures.
 Mineral oil has been found to prevent

evaporation from the culture and decrease the

metabolic rate of the culture by limiting the
supply of oxygen.
 This method is more suitable than lyophilization

for the preservation of non sporulating strains

 Culture Preservation
Lyophilization
 One of the best methods for long-term culture

preservation of many microorganisms is freeze-

drying (lyophilization).
 The commonly used cryoprotective agents are

skim milk (15% [wt/vol] for cultures grown on

agar slants and 20% for pelleted broth cultures)
or sucrose (12% [wt/vol] final concentration).
 It should be noted that some plasmid--

containing bacteria are successfully preserved by

this method.
 Culture Preservation
Liquid Drying
 To avoid the damage that freezing can cause, a

liquid—drying preservation process is applied.

 It has effectively preserved organisms such as

anaerobes that are damaged by or fail to survive

freezing.
 This procedure was preferred over lyophilization

for the maintenance of the biodegradation

capacity of six gram-negative bacteria capable of
degrading toluene.
 Culture Preservation
Cryopreservation
 Microorganisms may be preserved at - 5 to - 20°C for 1, to

2 years by freezing broth cultures or cell suspensions in

suitable vials.
 Deep freezing of microorganisms requires a

cryoprotectant such as glycerol or dimethyl sulfoxide

(DMSO) when stored at -70°C or in the liquid nitrogen at -
156 to -196°C.
 Broth cultures taken in the mid-logarithmic to late

logarithmic growth phase are mixed with an equal volume

of 10 to 20% (vol/vol) glycerol or 5 to 10% (vol/vol) DMSO.
 Alternatively, a 10% glycerol-sterile broth suspension of

growth from agar slants may be prepared.

 Culture Preservation
Protocol for Cryopreservation with Cryoprotectants by a Two-
stage Freezing Process, and Revival of Culture
 After centrifugation the supernatant is removed and the pellet,

consisting of microbial cells, is dissolved in an ice-cold solution

containing polyvinyl ethanol (10% [wt/vol]) and glycerol (10%
[wt/vol]) in a 1:1 ratio.
 Due to the presence of polyvinyl ethanol, a viscous thick cell

suspension is obtained, which is kept for about 30 minutes in an ice

bath for equilibration.
 During equilibration, an aliquot of 0.5 to 1.0 ml of the cell

suspension is dispensed into each plastic cryovial or glass ampoule.

 They are tightly closed, clamped onto labeled aluminum canes, and

placed at -30°C for about 1 h or for a few minutes in the gas phase of
liquid nitrogen to achieve a freezing rate of about 1°C/min.
 The canes are then placed into canisters, racks, or drawers and

frozen rapidly at -80°C or in liquid nitrogen.

 Culture Preservation
Preservation in Liquid Nitrogen
 Storage in liquid nitrogen is clearly the preferred

method for preservation of culture viability

Improvement of Saccharomyces yeast
strains for brewing, wine and baking
 Introduction
 Yeast is widely distributed in the nature, fruit
skins .
 Unicellular fungi species
 Around 350 species (39 genera) are accepted as
yeasts.
 Used for brewing, wine making, distilling and
baking.
 Commercials include Saccharomyces, Candida
and kluyveromyces.
 The use of starter cultures in yeast industries
became a common practice after methods for the
isolation of pure yeast strains were developed.
 Introduction
 Moreover, effort has been undertaken to improve
these strains, first by classical genetic methods
and later by genetic engineering.
 In general, yeast strain development has aimed at

improving the velocity and efficiency of the

respective production process and the quality of
the final products.
Industrial processes
 Industrial processes
1. Baker’s yeast
 One of the largest fermentation industry
 Around 1.8 million/year produced
 Four active forms are available

1) Compressed, 2) Cream, 3) Active dry 4) Instant

active dry yeast.
 Production of baker’s yeast
Yeast strain used
 Non-sproulating ‘torula’ yeasts have occasionally

been used for baking; specially selected strains of

Saccharomyce cerevisiae are used.
 For some time two strains of baker’s yeasts were

available: one was highly active but had poor but

had poor stability during storage ; the other had
poor activity but was highly stable in storage.
 Successful breeding program were then
undertaken to produce new strains from them.
Industrial processes
Yeast strains used for the modern fast-rising
dough have been developed with the following
traditional and new physiological properties in
mind:-
1. ability to grow rapidly at room temperature of
about 20-25oC ;
2. easy dispensability in water
3. ability to produce large amount of CO2 in flour
dough, rather than alcohol;
4. good keeping quality, i.e., ability to resist
autolysis when stored at 20oC;
 Industrial processes
5. high potential glycolytic activity ;
6. ability to adapt rapidly to changing substrates;
7. high invertase and other enzyme activity to
hydrolyze the higher glucofructans rapidly;
8. ability to grow and synthesize enzymes and
coenzymes under the anaerobic conditions of the
dough;
9. ability to resist the osmotic effect of salts and
sugars in the dough;
10. high competitiveness i.e high yielding in terms
of dry weight per unit of substrate used.
 Industrial processes
Culture maintenance
 The specially selected baking strains of

Saccharomyce cerevisiae are opt to mutate and

therefore proper storage is most important.
 Storage in liquid nitrogen and the oil culture

method in which sterile oil is placed over a slant

of yeast and refrigerated at 4oC are most widely
used.
 Freeze drying is not highly used as it induces

loss of viability in many yeasts and tendency

towards mutation.
 Industrial processes
Factory production
Substrate
 The substrate usually used for baker’ yeast

production is molasses.
 Any suitable sugar-containing substrate eg. Corn

steep liquor, sulphite Liquor.

 Beet and cane molasses , when they are

simultaneously available, are treated separately:

clarified, pH adjusted and sterilized.
 They are then mixed in equal amounts so that the

nutritional deficiency of one type is made up by

the other.
Industrial processes
 Cane sugar is particularly richer in biotin,
panthothenic acid, thiamin and magnesium and
calcium; while beet molasses is much richer in
nitrogen.
 When only one type of molasses is available,

deficiencies are made up by adding appropriate

nutrients.
 Clarification of molasses are required to remove

inert color material and also help to reduce foaming.

 Sterilization is achieved by heating at 100o-110oC

for about an hour, after the pH has been adjusted to

pH 6-8 to prevent caramelization of the sugar.
 Industrial processes
Fermentor process
 The fermentor for baker’s yeast propagation is

nowadays made of stainless steel.

 The trade fermentor may be anything from 75 to 225

cubic meters.
 Of this about 75% is occupied by the medium, the

unused space being allowed for foaming.

 The typical stirred tank fermentor with agitator

baffles and sparging is not often used in yeast

growth because of the high initial and operating cost.
 Continuous fermentation has not been widely used

in baker’s yeast production.

 Industrial processes
Harvesting the yeast
 The period of fermentation in the trade or

production fermentor varies from 10 to 20 hours

depending how much yeast is pitched into it; cells
form from 3.5% to 5% dry wet of the broth.
 The fermentation broth is cooled and cells

concentrated in centrifugal separators; they are

washed by resuspension in water and
centrifugation until they are light in color.
 Industrial processes
Baker’s yeast form
1. Compressed yeast: the yeast product obtained
after harvesting, is mixed with fine particles of ice,
starch, fungal inhibitors and processed vegetable
oil which help to stabilize it.
2. Active dry yeast: dry yeast is more stable in that
it can be used in areas or countries where
refrigeration is not available.
In many developing countries baker’s yeast is
imported from abroad in the form of active dry
yeast.
 Industrial processes
2. Food yeasts
 Yeasts are used for food by man for the following

reasons: to provide protein; to impart flavor and

to supply vitamins especially B-vitamins.
 Food yeasts are sometimes prescribed medically

when a deficiency of B-vitamins exists in a

patient.
 Industrial processes
Production of Food yeasts
 Food yeasts are produced from a wide variety of

yeasts and substrates.

 Yeasts used as food yeasts: Sacch. cerevisiae,S.

carlbergiensis, S. fragilis, Candida utilis, and

Candida tropicalis.
Substrates used for food yeast production
 The most commonly used substrates are molasses,

sulphite liquor, wood hydrolysate, and whey.

 Since interest developed in single cell protien other

unconventional sources have been developed.

 These include hydrocarbons, alcohol and wastes of

various types.
 Industrial processes
 The use of continuous fermentation was attractive
because the sulfite is produced almost continuously in
the operation of the pulp factory.
 In general a waldhof-type fermentor is used for the
continuous production of yeasts from sulfite waste.
 Liquors from various sources are usually blended.
 Thereafter, the sulfite containing compounds are
removed either by precipitation with lime, by aeration
by passing steam through it.
 The pH is adjusted from about 2 to 5.5 using ammonia.
 The lowest pH consistent with high yield is usually
preferred in order to lessen the chances of
contamination.
Industrial processes
3. Alcohol Yeasts
 Alchol yests are those to be used in beer brewing

wineries, and distilleries for spirits of industrial

alcohol.
 In the production of alcohol yeasts, the aim is cell

production.
 The methods are generally similar to those

already described for baker’s yeasts.

 Beginning from a lypholized vial or tube,

contamination is checked in a plate. A single

colony is picked and multiplied in sequentially
increasing amounts.
 Industrial processes
 The yeasts used are specially selected strains of the
following:
 Brewing: S.cerevisiae, S. uvarum carlbergensis S.
uvarum.
 Wine: S. cerevisiae, S. bayanus, S.beticus, S.
elipsoides.
 The medium used in the multiplication of the yeast is
made of materials to be found in the final
fermentation.
 Thus for growing brewery yeasts wort is used, for
distiller’s yeast a rye-malt medium is used, and for
wine grape juice is used.
 Alcohol yeasts are usually recovered and reused for
several rounds of fermentation before being discarded.
Industrial processes
Industrial processes
Industrial processes
Industrial processes
Yeast products
 Various products used in the food,

pharmaceutical and related industries may be

produced from yeasts.
 Yeast extracts are used in the preparation of

soups, sausages, gravies, to which they impart

a meaty flavor.
Industrial processes
 The yeast extract may be obtained by autolysing
the yeasts and thereafter spray-dring or drum-
drying with or without extracting soluble
materials from the autolysate.
 The extract may also be obtained by hydrolyzing

the yeast cells in acid solution.

 It is neutralized with sodium hydroxide, filtered,

decolorized through charcoal and concentrated

to syrup or spray dried.
 Yeast products are usually fortified with the

flavoring compound, mono-sodium glutamate,

extracts of animal or vegetable protein or with
yeast cells.
 Industrial processes
 Yeast extract are consumed for dietary purpose on
pharmacetical grounds as a source of vitamins, mainly
vitamin B12.
Future Developments in Alcoholic Beverage
Fermentations
 During alcoholic beverage production, many

developments take place involving the yeasts

strains employed in fermentations.
 Research is aimed not only at improving the

efficiency of sugar conversion to alcohol, but also in

selecting new yeasts to impart desirable flavours
and to utilize different carbohydrate raw materials.
 For Scotch whisky production, have identified

desirable characteristics for distilling yeast strains.

 Industrial processes
In brewing and winemaking , rapid advances in molecular
biology have influenced the application of novel yeast
strains in wort and grape juice fermentations, respectively.
These developments, notably recombinant DNA

technology, can lead to improved fermentation

performance and final product quality .
 Yeast-yeast genetic modification, also called self-cloning,

represents an attractive technique due to more favourable

regulatory issues and consumer acceptability .
 It is important to note, however, that genetically-modified

(GM) S. cerevisiae strains for food and beverage

fermentations have so far not been widely adopted and are
only authorized for use in some countries, but not in
Europe .
