Bioengineering in-production-of-food-ingredients

OMICSGroupeBooks
001
Progress in
Biotechnology for
Food Applications
www.esciencecentral.org/ebooks
Edited by
Dr. Wing-Fu Lai

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Progress in Biotechnology for Food Applications
Chapter: Bioengineering in Production of Food Ingredients
Edited by: Wing-Fu Lai
Published Date: June 2014
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Abstract
Bio/Genetic engineering allows genetic material to be transferred between any two
organisms, including between plants and animals. For example, the gene from a fish that lives
in very cold seas has been inserted into a strawberry, allowing the fruit to be frost-tolerant.
However, this has not yet been done for currently available commercial food crops. Concerns
about climatic change may lead to increased development and use of drought tolerant GM food
crops. Comparison to the corresponding unmodified organism essentially regulates products
of biotechnology on a par with organisms produced by traditional methods. This type of
comparison equilibrates the new and old risks by removing new risks commensurate with old,
unregulated risks from the regulatory process. However, because it requires a judgment by
the manufacturer involving risks that may be unknown and unquantifiable, environmentalists
and others skeptical of the new technology may feel that it leaves too many products of that
technology unregulated or under regulated. Critics may also think that manufacturers using
the new technology have too much discretion to unilaterally decide whether their products
meet the required standards.
Keywords: Bioengineering; Food ingredients; Genetically modified foods
Introduction
Food technology is a branch of food science in which modern biotechnological methods
are applied to improve food production or food itself. Various biotechnological processes
used to create and improve new food and beverages product include industrial fermentation,
plant cultures and genetic engineering. Food biotechnology was advanced in 1871 when
Louis Pasteur discovered that heating juices to a certain temperature would destroy bad
bacteria which would affect wine and fermentation. This process was then applied to milk
production, heating milk to certain temperature to improve food hygiene. Food science and
food biotechnology was then progressed to include the discovery of enzymes and their role in
fermentation and digestion of foods. With this invention further technological development of
enzymes emerged. Typical industrial enzymes used plant and animal extracts. Later this was
substituted by microbial enzymes. Foods genetically modified using biotechnology is known as
GM foods. Genetic material is altered using non-traditional, laboratory based methods; this is
Omprakash H Nautiyal*
Professor of Organic Chemistry/Natural Products Chemistry, Vadodara, Gujarat, India
*Corresponding author: Omprakash H Nautiyal, Professor of Organic Chemistry/Natural
Products Chemistry, 102, Shubh Building, Shivalik II, Canal Road, Chhani Jakat Naka, Vadodara
390002, Gujarat, India, Tel: +91-8733974519; E-mail: opnautiyalus@yahoo.com
Bioengineering in Production of Food
Ingredients

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known as genetic engineering. Individual genes with speficific desirable traits are transferred
from one organism to another [1].
An example of this development would be the use of chymosine in the production of cheese
which was typically made using the enzyme rennet and was extracted from the stomach lining
of the cow. Scientists then started using a recombinant chymosine in order for milk clotting
resulting in cheese curds. Food enzyme production using microbial enzymes was the first
application of the genetically modified organisms. Food biotechnology has grown to include
cloning of plants and animals as well as more developed genetically modified foods in more
recent years. In 1946 scientists discovered that DNA can transfer between organisms. The first
genetically modified plant was produced in 1983, using an antibiotic resistant tobacco plant.
In 1994 the transgenic flavr Savr tomato was approved by the FDA for marketing in the US.
The modification allowed the tomato to delay the ripening after picking.
Traditional breeding can achieve similar effects, but works over a much longer time span
and is not as targeted as GM. In addition, traditional breeding cannot transfer genes from
unrelated species as is possible with GM foods. Genetic modification of food is not new. Humans
have been altering food crops and animals through selective breeding for many centuries.
However, while genes can be transferred during selective breeding, the scope of exchanging
genetic material is much wider using genetic engineering [2].
In theory, genetic engineering allows genetic material to be transferred between any two
about climatic change may lead to increased development and use of drought tolerant GM food
crops [3,4].
Some foods and fiber crops have been modified to make them resistant to insects and
viruses and more able to tolerate herbicides. The major crops that have been modified for these
purposes, with approval from the relevant authorities, are [5,6]:
• Maize (corn)
• Wheat
• Rice
• Oilseed rape (canola)
• Chicory
• Squash
• Potato
• Soybean
• Alfalfa
• Cotton
Modified genes are being used in whole foods such as wheat, soybeans, maize and
tomatoes. These GM whole foods are not presently available in Australia. GM food ingredients
are however present in some Australians food. For example soy flour in bread may have come
from imported GM soybeans [7,8].

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Modified genes may have been used in an early stage of the food chain, but may or may not
be present in the end product. Nevertheless, gene products for example, phytochemicals (plant
chemicals that contain compounds which may prevent disease) may remain in the food chain.
The implications for human health are unknown.
Foods certified as Organic or biodynamic should not contain any GM ingredients according
to industry guidelines.
Inexpensive, safe and nutritious foods are needed to feed the world’s growing population.
Genetic modification may provide:
• Sturdy plants able to withstand weather extremes.
• Drought-tolerant and salt-tolerant crops.
• Better-quality food crops.
• Higher nutritional yields.
• Inexpensive and nutritious food, such as carrots with more antioxidants.
• Foods with a longer shelf life, like tomatoes that taste better and last longer.
• Food with medicinal (nutraceutical) benefits, such as edible vaccines – for example,
bananas with bacterial or rotavirus antigens.
• Disease and insect resistant crops that require less pesticide and herbicide – for
example, GM canola [9-11].
GM advocates argue that GM foods are better for the environment. By using GM crops
that are resistant to attack by pests or disease, farmers can reduce their use of pesticides and
herbicides and the residual levels of these chemicals in the environment. However, development
of resistance can undermine and even reverse this benefit [12,13].
Genetic engineering can be used to increase amounts of particular nutrients (like vitamins)
in food crops. Research into this technique, sometimes called nutritional enhancement is now
at an advanced stage.
GM golden rice is a white rice crop modified by the insertion of the vitamin A gene from
a daffodil plant. This changes the color and the vitamin level of the rice and is of benefit in
countries where vitamin A deficiency is prevalent.
GM researchers are focusing on major health problems like iron deficiency. The removal
of the proteins that cause allergies from nuts (such as peanuts and Brazil nuts) is also being
studied [14,15].
The Risks of GM Crops
Concerns about genetic modification of food raised by scientists, community groups and
members of the public include:
New allergens could be inadvertently created – known allergens could be transferred from
traditional foods into GM foods. For instance, during laboratory testing, a gene from the Brazil
nut was introduced into soybeans. It was found that people with allergies to Brazil nuts could
also be allergic to soybeans that had been genetically modified in this way and so the project
was ceased. No allergic effects have been found with currently approved GM foods [16,17].

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Antibiotic resistance may develop – bioengineers sometimes insert a marker gene to help
them identify whether a new gene has been successfully introduced to the host DNA. One such
marker gene is for resistance to particular antibiotics. If genes coded for antibiotic resistance
enter the food chain and are taken up by human gut micro flora, the effectiveness of antibiotics
could be reduced and human infectious disease risk increased. Research has shown that the
risk is very low; however, there is general agreement that use of these markers should be
phased out [18-20].
Cross-breeding – GM crops can cross-breed with surrounding vegetation, including weeds,
transferring undesired characteristics. The introduction of glyphosate-resistant soybeans in
1996 has produced glyphosate-tolerant weeds that have driven even greater use of herbicides.
Pesticide-resistant insects – the genetic modification of some crops to produce the natural
biopesticide Bacillus thuringiensis (Bt) toxin could encourage the evolution of Bt-resistant
insects, rendering the spray ineffective.
Biodiversity – growing GM crops on a large scale may affect the balance of wildlife and the
environment. Since bees cannot distinguish GM from non-GM crops, GM crops can affect non-
GM and organically–farmed crops through cross-pollination.
Cross-contamination – plants bioengineered to produce pharmaceuticals (such as
medicines) may contaminate food crops.
Health effects – minimal research has been conducted into the potential acute or chronic
health risks of using GM foods.
Social and Ethical Concerns about GM
Concerns about the social and ethical issues surrounding genetic modification include:
The possible monopolization of the world food market by large multinational companies
that control the distribution of GM seeds
Concerns related to using genes from animals in plant foods. For example, eating traces of
genetic material from pork is problematic for certain religious and cultural groups
Animal welfare could be adversely affected. For example, cows given more potent GM growth
hormones could suffer from health problems related to growth or metabolism
New GM organisms could be patented so that life itself could become commercial property
[21,22].
Regulation of GM foods
In Australia, GM foods are regulated by the Food Standards Australia New Zealand (FSANZ)
Code under Standard 1.5.2 – Food produced using Gene Technology. GM foods receive
individual pre- market safety assessments prior to use in foods for human consumption.
A GM food will only be approved for sale if it is assessed as being safe and as nutritious as
its conventional counterparts. The assessment investigates;
• Nutritional content
• Toxicity (using similar methods to those used for conventional foods)
• Tendency to provoke any allergic reaction

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• Stability of the inserted gene
• Whether there is any nutritional deficit or change in the GM food
• Any other unintended effects of the gene insertion
The safety of GM foods is still being debated, as it is impossible to predict all of the potential
effects on human health and the environment. Consequently, some public health experts
advocate caution, believing that we do not know whether GM foods are safe [23,24].
GM labeling
Since December 2002, Australia law has required that food labels must show if food has
been genetically modified or contains GM ingredients, or whether GM additives or processing
aids remain in the final food product. The label on the package must include the statement
‘genetically modified’ in conjunction with the name of the food or ingredient or processing aid.
GM foods are labeled to help consumers make informed decisions about the food they buy,
not for safety reasons. Special labels are not required for:
‘Highly refined’ foods that no longer contain the altered DNA or protein (for example, oil
from modified corn)
GM food additives or processing aids (unless the new DNA remains in the food to which it
is added)
GM flavors constituting less than 0.1 per cent of the food by weight
Food, food ingredients or processing aids unintentionally containing less than one per cent
GM material
Food prepared at point of sale (takeaway and restaurant food does not have to be labeled).
Labels may be required if:
Genetic modification has altered the food so that its composition or nutritional value is
outside the normal range of similar non-GM goods (for example, high omega-3 soybeans)
Food contains toxins which are significantly different to those in similar non-GM foods
The food produced using GM technology contains a new factor which can cause allergic
reactions in some people
Genetic modification raises significant ethical, cultural and religious concerns regarding
the origin of the genetic material used [25-27].
GM food on the shelves
Many foods on supermarket shelves contain imported GM ingredients. GM foods have also
been approved for production in Australia, including corn, soybeans, potatoes, canola and
rice.
Other GM foods are still undergoing field trials approved by the Office of the Gene Technology
Regulator, although the moratorium by state governments (lifted in Victoria and NSW in early
2008) stopped some trials. Imported food products are subject to the same regulations as
domestically manufactured foods.
Around 20 GM foods, additives, flavorings, growth hormone (bovine somatotropin) and
enzymes (like rennet, used to make cheese) are currently approved in Europe. More than 40
GM foods are approved for sale in the USA [28,29].

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The Main Sources of GM Foods in Australia
Soya imported from the United States – the soya has been genetically modified to be
resistant to herbicide. It can be found in a wide range of foods, such as chocolates, potato
chips, margarine, mayonnaise, biscuits and bread.
Cottonseed oil made from GM cotton – this oil, made from cotton resistant to a pesticide,
is used in Australia for frying (by the food industry) and in mayonnaise and salad dressings.
Imported GM corn – this is mainly used as cattle feed at present and has not been approved
for farming in Australia. However, GM corn may have entered the Australian market through
imported foods like breakfast cereal, bread, corn chips and gravy mixes. If so, it is now required
to be labeled.
GM ingredients in imported foods including GM potatoes, canola oil, rice, sugar beet, yeast,
cauliflower and coffee.
Foods genetically modified using biotechnology is known as GM foods. Genetic material is
altered using non-traditional, laboratory-based methods; this is known as genetic engineering.
Individual genes with specific desirable traits are transferred from one organism to another
[30,31].
Traditional breeding can achieve similar effects, but works over a much longer time span
and is not as targeted as GM. In addition, traditional breeding cannot transfer genes from
unrelated species as is possible with GM foods.
Genetic modification of plants and animals
Genetic modification of food is not new. Humans have been altering food crops and animals
through selective breeding for many centuries. However, while genes can be transferred during
selective breeding, the scope for exchanging genetic material is much wider using genetic
engineering.
In theory, genetic engineering allows genetic material to be transferred between any two
about climate change may lead to increased development and use of drought-tolerant GM food
crops [32,33].
The Society of Toxicology (SOT) is committed to protecting and enhancing human, animal,
and environmental health through the sound application of the fundamental principles of
the science of toxicology. It is with this goal in mind that the SOT defines here its current
consensus position on the safety of foods produced through biotechnology. In this context,
biotechnology is taken to mean those processes whereby genes that are not endogenous to
the organism (transgenes) are transferred to microorganisms, plants, or animals employed in
food production, or where the expression of existing genes is permanently modified, using the
techniques of genetic engineering. We intentionally avoid using the term Genetically Modified
Organisms (GMOs) or foods in this context, since conventional techniques of plant and animal
breeding, which are not considered here, also involve genetic modification. The extent of the
genetic changes resulting from such conventional breeding techniques, which is generally
undefined, far exceeds that typically produced by transgenic methods. Consequently, it is
important to recognize that it is the product, and not the process of modification, that is the

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focus of concern regarding the human or environmental safety of Biotechnology-Derived (BD)
foods [34,35].
The principal responsibilities of toxicologists are to define and characterize the potential
for natural and manufactured materials to cause adverse health effects and to assess, as
accurately as possible, the plausibility and level of risk for human or animal health or for
environmental damage under a defined set of circumstances. It is not the task of the Society
of Toxicology to determine the overall value of a product or process by balancing health or
environmental risks with potential benefits, or to choose between different strategies to manage
risk, although toxicological considerations are important in both processes. Our purpose here
is rather to identify and consider the primary toxicological issues associated with BD foods.
Major areas of concern in the development and application of such foods in agriculture relate
to the possibility of deleterious effects on both human health and the environment. We do not
consider here some aspects of the possible environmental impact of GM organisms such as
gene transfer to non-engineered plants [36,37].
Current techniques of developing organisms used in the production of BD foods typically
involve the transfer to the host of the desired gene or genes in combination with a promoter
and a gene for a selectable marker trait that allows the efficient isolation of cells or organisms
that have been transformed from those that have not. Common selectable markers in plants
have included resistance to antibiotics (kanamycin/neomycin or ampicillin) or herbicides [38].
Several key issues have been raised with respect to the potential toxicity associated with
BD foods, including the inherent toxicity of the transgenes and their products, and unintended
(pleiotropic or mutagenic) effects resulting from the insertion of the new genetic material into
the host genome. Unintended effects of gene insertion might include an over-expression by the
host of inherently toxic or pharmacologically active substances, silencing of normal host genes,
or alterations in host metabolic pathways. It is important to recognize that, with the exception
of the introduction of marker genes, the process of genetic engineering does not, in itself,
create new types of risk. Most of the hazards listed above are also inherent in conventional
breeding methods [39].
It has been 17 years since the groundbreaking 1975 meeting at A silo mar where scientists
discussed the emerging technology of molecular biology, its vast potential and the possible
risks that could result from the ability to transfer DNA from one organism to another.1 Since
then, a number of biotechnology-derived pharmaceutical products have already gone on the
market,2 and the first food and agricultural products have been approved or are close to
approval.3 Many more such products are under development, and there have been no adverse
impacts on human health or the environment. Rather, reputable scientific and medical sources
stress the potential of biotechnology to improve human health and nutrition, and to ameliorate
the adverse impacts of traditional agricultural practices on the environment. Despite the
abundance of data indicating the beneficial potential of biotechnology and the absence of
harmful incidents, genetic engineering has aroused considerable public suspicion and from
some quarters a demand for government oversight out of proportion to the demonstrated risks.
The negative perception and resulting regulatory response threatens to adversely affect the
development and competitiveness of this fledgling industry, and may also delay or even block
the introduction of beneficial products [40].
This Comment examines issues in the regulatory oversight of the production and
consumption of bioengineered food. As an introduction, the Comment first examines the

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range of products under development and their potential benefits and risks. It next considers
the recent controversy over the use of genetically engineered bovine growth hormone, which
illustrates many of the issues in this area. The next section presents biotechnology’s possible
risks and then discusses the advantages of a comparative approach to risk regulation of
bioengineered food. Next, the Comment examines the current government regulation of this
technology. Since the potential risks involve both the genetically engineered food products
and the environmental release of the organisms that produce them, regulation in these areas
is analyzed. This regulation has been implemented through the “coordinated framework,” an
adaptation of existing statutes to the oversight of biotechnology. Although regulation under
the coordinated framework has the advantage of not singling out biotechnology for special
oversight, the Comment examines some of the criticisms that stem from adapting existing
statutes to address biotechnology. The Comment next considers, in light of the benefits
of a comparative approach, some recently proposed approaches to regulation. Finally, the
Comment contemplates what modifications in government policy and regulation might improve
the public perception of biotechnology, strengthen the industry, and foster the generation of
products that are not merely profitable, but also truly beneficial to both human health and the
environment [41].
This Comment examines issues in the regulatory oversight of the production and
consumption of bioengineered food. As an introduction, the Comment first examines the
range of products under development and their potential benefits and risks. It next considers
the recent controversy over the use of genetically engineered bovine growth hormone, which
illustrates many of the issues in this area. The next section presents biotechnology’s possible
risks and then discusses the advantages of a comparative approach to risk regulation of
bioengineered food. Next, the Comment examines the current government regulation of this
technology. Since the potential risks involve both the genetically engineered food products
and the environmental release of the organisms that produce them, regulation in these areas
is analyzed. This regulation has been implemented through the “coordinated framework,” an
adaptation of existing statutes to the oversight of biotechnology. Although regulation under
the coordinated framework has the advantage of not singling out biotechnology for special
oversight, the Comment examines some of the criticisms that stem from adapting existing
statutes to address biotechnology. The Comment next considers, in light of the benefits
of a comparative approach, some recently proposed approaches to regulation. Finally, the
Comment contemplates what modifications in government policy and regulation might improve
the public perception of biotechnology, strengthen the industry, and foster the generation of
products that are not merely profitable, but also truly beneficial to both human health and the
environment [42].
Biotechnology used in the Production of Food
Biotechnology as applied to the production of food (Figure1) has the potential to greatly
benefit the public, as well as to improve agricultural productivity. The Council on Scientific
Affairs of the American Medical Association has stated that agricultural biotechnology has the
potential to “meet the needs of a rapidly growing population and minimize the toxic influences
of traditional farming practices on the environment.” However, due to agricultural economics
as well as scientific complexity, the function of many of the first genetically engineered food
products is to improve agricultural efficiency and productivity. Since the technology for
producing foreign proteins in genetically engineered bacteria is more established than the

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technology for transforming entire plants or animals, some of the first products are proteins
that can be inexpensively produced to replace or augment the same naturally occurring
protein. Thus, the first genetically engineered food ingredient approved by the Food and Drug
Administration (FDA) is chymosin, an enzyme traditionally obtained from the stomach of
calves and used in the production of cheese. Since the genetically engineered chymosin is
identical to the enzyme obtained from the traditional preparation and contains no ingredients
that are not Generally Recognized As Safe (GRAS), the FDA has concluded that this product,
like the traditional product, is GRAS.8 Bovine somatotropin, a hormone used to increase milk
production, has been produced in genetically engineered bacteria and is virtually identical to
the naturally occurring protein. It is expected to be approved in 1992 [43].
Figure 1: Food production (http://www.ied.edu.hk/biotech/eng/classrm/class_food1.html).
Genetic engineering techniques have also been applied to commercial food crops. The “anti-
sense” tomato, one of the bioengineered products closest to FDA approval, has been modified
to retard the softening and subsequent spoilage that accompanies ripening. This modification
improves (Figure 2) the farmer’s ability to machine-pick ripe tomatoes without bruising them, thus
producing a tasty but easily harvested tomato. This type of modification improves the efficiency of
the agricultural industry and can result in increased supply and lower food prices for consumers.
Genetically-engineered herbicide-resistant plants that survive the application of herbicides
during weed eradication may also improve the efficiency of farming.12 Such plants will be used
in conjunction with recently developed herbicides that are rapidly biodegraded and are of low
toxicity.13 Finally, many crops, including tomato, tobacco, potato, alfalfa, cucumber, corn, and
soybeans have been genetically engineered to resist plant viruses that might otherwise devastate
these plants. This should improve both crop yield and quality, since harvested plants would have
far less viral contamination than is present in unmodified plants [44].

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Figure 2: Activation of silent genes. (http://www.naro.affrc.go.jp/org/nfri/
english/organization/bio/kouso.html National Food Research Institute).
The state of agricultural biotechnology today is that a variety of items are in development or
close to marketing. Those that are farthest along are of the type that will increase agricultural
efficiency or productivity, but the current technology has the potential for improving nutritional
quality and reducing the use of chemical pesticides. Moreover, there have been no hazardous
incidents that should create a fear of this technology and lead to tight regulatory oversight.
To the contrary, the Council on Scientific Affairs of the American Medical Association has
recommended that physicians play a role in educating the public that “genetic manipulation
is not inherently hazardous and that the health and economic benefits of recombinant DNA
technology greatly exceed any risk posed to society.”Yet public perception of biotechnology
is one of suspicion, leading to calls for tighter regulation. The following examination of the

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recent controversy over bovine somatotropin may illuminate some of these conflicting views on
biotechnology [45].
Bovine somatotropin
Bovine Somatotropin (bST), (Figure 3) also called bovine growth hormone, was the first
major product of recombinant DNA technology available for use in agriculture. bST occurs
naturally in cows, but when additional bST produced by genetically engineered bacteria is
administered to dairy cows, their milk production is expected to increase an average of 12%
without a commensurate increase in feed consumed. The use of bST has been heralded as
the technological advance that will have the most dramatic effect on the efficiency of milk
production in this decade. Moreover, numerous studies have shown that milk produced by
bST-dosed cows is safe for human consumption. The use of bST is also expected to lower
environmental pollution, because the decreased intake of feed relative to milk output will
decrease the production of manure, urine, and methane-a gas with a strong greenhouse
effect. Simply put, fewer cows are required to produce the same amount of milk. Despite these
benefits, bST has elicited considerable public outcry. Consumer groups are threatening to
boycott the milk from bST supplemented cows, grocery chains and food processing companies
refused the milk that was approved by the FDA for sale during the investigation period, and
states have considered or taken action either banning the use of bST or requiring labeling of
products derived from its use [46].
Figure 3: Production of bST and recombinant of DNA technology. (http://mmg-233-2013-genetics-
genomics.wikia.com/wiki/Bovine_somatotropin).
Though ten states introduced bills restricting the use of bST and two states actually
enacted such laws, only a few individuals generated the original controversy. Samuel Epstein,

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a professor at the University of Illinois, joined with genetic engineering critic Jeremy Rifkin to
attack the use of bST. In an evaluation of other scientific studies, Epstein questioned the safety
of bST-produced milk for human consumption; in addition, he concluded that bST adversely
affects the health of cows. Rifkin’s group, Foundation on Economic Trends (F.E.T.), demanded
the release of environmental assessment records relating to bST. F.E.T. also petitioned the
FDA for an environmental impact statement prior to field testing bST. The petition asserted
that use of bST would (1) significantly affect agricultural land use in milk-producing regions
of the United States; (2) adversely affect the internal environment of cattle injected with [bST];
and (3) have adverse economic and social impacts on the dairy industry [47].
The bST controversy illustrates some of the difficulties that may be encountered during the
introduction of bioengineered food products. Since biotechnology products are already suspect
in the public eye, they are easily attacked by a vocal minority. Even if they meet the current
regulatory standards, they are especially vulnerable to criticisms that they have adverse
environmental, economic, or social impacts that may result from modern dairy or agricultural
practices as a whole rather than just from biotechnology. State or local agencies may impose
additional regulation that could impede the development of the biotechnology industry and
delay advances that might actually be environmentally, economically, or socially beneficial.
The question, then, is what regulatory balance should be struck between the potential or
perceived risks of biotechnology and its unknown but almost certain benefits [48].
Risks and regulatory issues concerning bioengineered food
Although new and unknown technologies are often viewed with suspicion, some features
of biotechnology make it particularly susceptible to an exaggerated perception of risk. Public
concern may stem from scientists themselves, who initiated a moratorium on some aspects
of genetic engineering in 1974. While scientists have since grown comfortable with the
technology, the public perception of unreasonable risks lingers on. A recent survey found that
52% of the public “believes that genetically engineered products are at least somewhat likely
to represent a serious danger to people or the environment.” Biotechnology often suggests
the “Frankenstein image.” While the current technology generally changes but a single gene,
producing a relatively small modification, many people may believe that any interspecies
exchange of genetic information results in a dramatic change. Perhaps such views underlie
the finding that 24% of a group aware of biotechnology felt that creation of hybrid plants and
animals through genetic engineering is morally wrong. Another aspect of biotechnology that
invites public concern is the ability of living things to reproduce; thus any deleterious effects of
genetically engineered organisms have the ability to escape human control and self-perpetuate
[48,49].
This leads to a fear that although a deleterious result is unlikely, if it occurs, the outcome
could be a problem of substantial magnitude. Such fears of an unlikely but potentially
disastrous outcome could greatly hinder the progress of biotechnology. A majority of the public
would object to the use of genetically engineered organisms if the risk were unknown.46 Food
products of biotechnology generate their own specific concerns. Production of bioengineered
food usually involves not only a consideration of the safety of the food for human consumption,
but also the safety of environmental release of the altered plant. The public’s perception
of potential danger from food biotechnology is enhanced by its heightened awareness of
environmental damage from the introduction of exotic species and of health problems that are
manifested only decades after exposure to the causative agent. Yet many similar risks from food
stem from traditional agricultural and plant breeding practices that are essential to provide
sufficient food to the growing population or to assure the taste, quality, and convenience
that consumers and farmers have come to expect. Thus, society accepts environmental risks

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of pesticide use and dispersal of domesticated plants and animals within certain limits and
tolerates low levels of pesticide residues in food. Other risks from food are inherent in the
food itself. Food contains many naturally occurring toxicants and carcinogens that are nearly
unavoidable in the ordinary diet [48,49].
Biotechnology presents few risks beyond those already accepted in traditional foods. As to
their environmental risks, “crops modified by molecular and cellular methods should pose risks
no different from those modified by classical genetic methods for similar traits.” Bioengineered
organisms’ potential for dispersal and environmental disruption is generally similar to their
traditional counterparts. Society has long accepted the fact that traditional plant and animal
breeding practices may change the nutrient or toxicant levels in the food or alter an organism’s
potential for environmental dispersal. Although traditional methods usually enhance the safety
of the food, they have occasionally increased the level of a deleterious component. The use of
antibiotic resistance marker genes in the production of bioengineered food has raised some
questions, but most experts agree that the genes should cause no health or safety problem.
Bioengineering as an extension of traditional breeding practices, should pose no greater
concern over the safety of the food consumed; it should actually be safer since the recombinant
techniques are more specific and thus less likely to produce unwanted side effects such as
increased levels of toxicants or weediness. Indeed, as considered above, bioengineering may
lower both the environmental and food consumption risks [48,49].
Thus, to foster technological advance and its resultant benefits, Huber argues that a
comparative system of regulation of old and new risks, one that permits new technologies
functionally similar to established technologies and of no greater risk, should be implemented.
The comparative approach, allowing a new risk, is justified when the old, risky product is one
that society accepts either because it is essential or desirable. “Excessively strict regulation
of the safer-than-average products will drive consumption toward the more hazardous ones.”
Comparative regulations, on the other hand, would favor the safer product, particularly
because modern technology usually replaces an old outmoded source of risk rather than
adding to it [48,49].
Huber suggests a four-step process for implementing comparative regulation:
1) The agency must define a risk market comprising products that are functional substitutes
for each other.
2) It next must identify typically risky products already allowed to compete in that market.
3) The agency must then compare the risks of the new substitute with those of products
not in fixed supply and already in the market. Only the less safe substitutes must be excluded
or otherwise regulated.
4) If a new product offers exceptional price or other advantages over existing, more
hazardous products, introduction of the safer product could conceivably increase net risk
by increasing total consumption. As a final step in comparative regulation, an agency must
therefore consider whether a candidate for regulation is this type of risk.
The regulatory framework for bioengineered food is in transition from an approach that, by
focusing on the process used to produce genetically engineered food, did not always accurately
assess the risk of the product. The Bush Administration sought to cure this problem by
adopting a policy similar to the comparative regulatory approach discussed above. The federal
regulatory agencies are currently implementing this policy. This policy approach has the
advantage of removing unjustified oversight of biotechnology, but it may have the disadvantage
of under regulating the field, especially because it relies on existing statutory authority not
directed at biotechnology. Moreover, this policy approach does not address the non-risk based

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social and economic concerns that contribute to the public’s objections to biotechnology. It
may, therefore, fuel the demand for state and local regulation of biotechnology [48,49].
The regulatory framework
Regulation of bioengineered food falls under the general regulatory scheme that has been
established for biotechnology as a whole.
The “Coordinated Framework for the Regulation of Biotechnology” (coordinated framework),
introduced by the Office of Science and Technology Policy (OSTP) in 1985-86, describes the
policies for federal regulation of biotechnology. Under the coordinated framework, regulation
of biotechnology relies on existing federal statutes, with each agency maintaining jurisdiction
over biotechnology applications within its traditional domain. Oversight of each product is
within a single agency, but where more than one agency is involved, one is designated the lead
agency. Agencies rely upon existing statutory authority to provide immediate health and safety
protection, as well as to eliminate any regulatory delay or uncertainty that might hurt the new
biotechnology industry. Underlying this decision was the premise that genetic engineering
techniques are basically extensions of the traditional techniques of selective breeding and
hybridization, and thus the laws that governed products of those techniques could also apply
to biotechnology [48,49].
The Biotechnology Science Coordinating Committee (BSCC), established by the OSTP in
1985, had broad authority for promoting cooperation between the agencies and establishing
consistent scientific policy and reviews. It was composed of senior policy officials from the
United States Department of Agriculture (USDA), the FDA, the National Institutes of Health
(NIH), the Environmental Protection Agency (EPA), and the National Science Foundation (NSF).
In late 1990, the BSCC was replaced by the Biotechnology Research Subcommittee (BRS) of the
interagency Committee on Health and Life Sciences. The BRS is said to have responsibilities
similar to the BSCC. Although the BSCC, in its evaluation of the issues, could “develop generic
scientific recommendations that could be applied to similar, recurring applications,” it did not
re-evaluate agency decisions and thereby delay that agency’s response. Two important facets
of BSCC’s initial mission were to ensure that its constituent agencies regulate biotechnology
using scientific reviews of similar stringency, and to establish consistency as to which
genetically engineered organisms were subject to regulatory oversight [48,49].
The FDA has followed a policy, consistent with that stated in the coordinated framework,
that oversight of biotechnology products under its jurisdiction requires no new procedures
or requirements. The FDA is responsible for assuring the safety and quality of both plant
and animal bioengineered food products. New animal drugs, including those produced by
biotechnology, require pre market approval by the FDA. Moreover, the FDA must approve for
human consumption the edible portions of animals that have been administered a new drug. To
approve the use, the FDA must confirm the safety of the food product for human consumption;
the drug must not accumulate as unsafe residues in the edible portions of the animal. Finally,
the efficacy of the drug and its safety for both the animals and the environment must be
established. When a new drug produced by genetic engineering is virtually identical to an
approved substance produced by conventional technology, the showing for approval is reduced
and only a supplemental application to the FDA is necessary. Regulation of biotechnology-
derived foods from plants will depend largely on the use of the food. Generally, regulations
differ for “whole foods” such as fruits, vegetables, or grains; for substances unintentionally
added to foods; and for food additives. The FDA does not require pre market approval for

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whole foods, but the burden is on the producer or manufacturer to assure that such foods
are safe. However, the FDA can regulate whole foods, including those that are products of
biotechnology, under section 402(a)(1) of the FDCA, which sets different safety standards
for inherent natural constituents of the food and unintentionally added substances that are
poisonous or deleterious. Naturally occurring constituents posing safety problems, such as
elevated levels of solanine in a new potato variety or poisons in a toxic mushroom, make
the food legally adulterated only “if the quantity of such substance[s] ordinarily renders[s] it
injurious to health.” Unintentionally added substances on the other hand, are contaminants
and are subject to a more rigorous standard. The contaminants may be chemicals introduced
accidentally by human activities (e.g., Polychlorobiphenyls (PCBs), mercury, and lead) or they
may be naturally occurring contaminants (e.g., aflatoxin). “Added substance[s]” causes a food
to be legally adulterated if they “may render it injurious to health.” Adulterated food is subject
to an enforcement action if it enters into interstate commerce [48,49].
Food additives are subject to pre market clearance by the FDA, unless the additive is
Generally Recognized As Safe (GRAS). A substance is GRAS either if its safety is known from
common use in foods consumed by a significant number of consumers prior to January 1,
1958, or if its safety is determined by well-controlled scientific studies. A company may market
a product, believing it to be GRAS, but it runs the risk that the FDA may decide that it is not
GRAS and force it off the market. A company wishing to clarify the matter at the outset may
obtain the FDA’s opinion on the substance by filing a GRAS affirmation petition. If a substance
added to food is not GRAS, it is a food additive and under section 409 of the FDCA a company
must submit a food additive petition for FDA approval. Thus, if a bioengineered product is a food
additive, it requires submission of scientific data showing that it is safe under the conditions
for which it will be used. Moreover, under the requirements of NEPA, the manufacturer must
prepare an environmental assessment or an impact statement if the manufacturing process or
the use of the food additive will significantly affect the environment [48,49].
Regulation under the coordinated framework: Not surprisingly, regulation of
bioengineered food products, and biotechnology in general, is often viewed as too stringent by
biotechnology companies, as too lax by environmentalists, and as lacking a sound scientific
basis by academicians. Biotechnology companies often cite regulatory uncertainty as a
substantial concern in developing new products. In recent years, the main regulatory hurdle
that food biotechnology companies have had to face has involved the release of genetically
engineered organisms [50].
The companies acknowledge that APHIS’ handling of small field tests has worked well and
that many delays have been due to suits or to local restrictions on release. Thus, at the federal
level, industry concerns for the future stem from uncertainty over regulation of large scale
release of genetically engineered crops during commercialization; the adequacy of coordination
between the EPA, the USDA, and the FDA when a single product requires oversight by all
three agencies; and over how the FDA will handle food products. Industry may actually
welcome a case-by-case analysis of the first bioengineered foods, because FDA approval will
give an assurance of safety that will boost public confidence in the products. In the long
run, however, industry representatives feel that bioengineered foods should require no more
screening than traditionally produced foods. Since under current law this would mean that
genetically engineered foods classified as whole foods would require no pre market approval,
the International Food Biotechnology Council, an industry association, recommends that
the FDA establish a voluntary pre market notification system. However, industry’s greatest

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regulatory concern may not be with federal regulations but with the increasing patchwork
of state and local regulations. This concern, which will be considered further below, has led
industry to lobby for more explicit federal regulation [50].
Environmental groups have faulted the coordinated framework for incompletely regulating
biotechnology through existing statutes not directed at genetic engineering and for not keeping
environmental considerations paramount. Environmentalists have criticized the use of the
Federal Plant Pest Act to regulate environmental release of genetically engineered agricultural
plants and animals because it covers only plant pests. Although use of the Ti plasmid as a
vector has brought most plant genetic engineering under the authority of the regulations,
environmentalists are concerned that increased use of other means of introducing foreign
DNA will leave many bioengineered plants unregulated. Moreover, environmentalists and
university researchers are concerned that USDA’s statutory authority is inadequate to cover
genetically engineered animals. Finally, environmentalists argue that the EPA, rather than the
USDA, should be the lead agency in charge of environmental release of genetically engineered
organisms, since EPA’s mandate is to protect the environment as a whole, whereas USDA’s
interest is to promote agriculture [50].
Specific policy changes reflecting this philosophy have been proposed both for the regulation
of the release of bioengineered food producing organisms and the assessment of food safety
and quality. The new policy concerning the release of food producing organisms mandates
equivalent, risk-based regulation of traditional and genetically engineered organisms, and thus
should lower industries’ burden of federal regulation. The new policy no longer specifies that
“inter generic organisms” or “pathogenic” species require oversight. The policy broadly covers
all types of genetic modifications, including those resulting from traditional methods, by stating
that “a determination to exercise oversight should not turn on the fact that an organism has
been modified by a particular process or technique.” Rather, all organisms should be regulated
according to the risk of introducing them into a particular environment. Federal agencies
should not exercise oversight of such introductions unless the risk is unreasonable. However,
federal agencies need not choose between imposing or not imposing oversight. Agencies have a
range of options, such as “issuance of suggested industry practices, development of guidelines
for certain introductions, and requirements for notification, labeling, prior review or approval
of certain introductions.” In determining what level of oversight should be applied, the policy
adopts a comparative approach similar to Huber’s. Thus, “an introduction should be subject
to no greater degree of oversight than was a comparable organism or product previously used
in past safe introductions in a comparable target environment.” In short, this policy suggests
a comparative approach to regulation by applying comparable oversight to comparable
organisms in similar environments. Such an analysis would apply whether the new organism
is, for example, a genetically engineered variety or a newly introduced exotic species [50,51].
One feature of this assessment, in addition to qualitative and analytical evaluation, is
a comparative approach. Acceptable and unacceptable levels of certain food constituents
are determined by reference to the host plant when that plant has a history of safe use. By
following FDA’s guidelines, a manufacturer is expected to determine whether the product is
safe, requires FDA consultation because of questionable safety, or is unsafe. Thus, the FDA
suggests that the level of toxicants in the new variety should be within the range of toxicant
levels in the host variety, and that “the concentration and bioavailability of important nutrients
in the new variety should be within the range ordinarily seen in the host species.” If toxicant
levels present a safety concern, the food is unacceptable; if nutrient levels are outside of the

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normal range, the manufacturer must consult the FDA to determine its course of action. The
primary concern raised by the donor species is the potential transfer of allergens or toxicants
to the host. The manufacturer must consult the FDA if it is possible that allergens have
been transferred from the donor to the host plant. The assessment of food safety may entail
qualitative, as well as quantitative comparisons. Thus, a manufacturer must consult the FDA
if the introduced protein, carbohydrate, fat, or oil is likely to be a major component of the diet
and is not derived from an edible source, or differs substantially from that in the edible source
[50,51].
Comparison to the corresponding unmodified organism essentially regulates products
of biotechnology on a par with organisms produced by traditional methods. This type of
comparison equilibrates the new and old risks by removing new risks commensurate with old,
unregulated risks from the regulatory process. However, because it requires a judgment by
the manufacturer involving risks that may be unknown and unquantifiable, environmentalists
and others skeptical of the new technology may feel that it leaves too many products of that
technology unregulated or under regulated. Critics may also think that manufacturers using
the new technology have too much discretion to unilaterally decide whether their products
meet the required standards. Moreover, it is difficult to see how agencies can apply a risk-
based policy to certain organisms (e.g., transgenic fish) perceived to be immune to oversight
due to gaps in statutory authority. Nor does the policy address ideological deficiencies that
result from using existing statutory and regulatory authority. Thus, while the policy may ease
regulatory burdens at the federal level, it may create a backlash from the public and from state
and local bodies that perceive greater risks from biotechnology and wish to regulate it more
stringently [50,51].
Horizontal Gene Transfer
Horizontal gene transfer (HGT) is the process by which an organism transfer genetic
material to another organism other than its offspring and which is followed by integration
and expression of the genetic material. This process is common among bacteria and other
prokaryotes. Speculation that HGT could occur between ingested bioengineered food and
enteric bacteria present in the human mouth, stomach, and gut has been expressed. Of
special concern are bioengineered foods made from transgenic plants that express antibiotic-
resistance markers (ARMs), which are employed during the development of the transgenic
plant to select for those that have incorporated the transgenes. When humans ingest food
derived from plants that express an ARM, it is theoretically possible that the ARM could be
taken up and stably integrated into enteric bacteria through HGT, resulting in bacteria that
are resistant to specific antibiotics. This situation has never been reported, although studies
point to its possibility. The epsps transgene, which confers resistance to a common herbicide,
survives intact through the small intestine of humans when bioengineered food made with
Roundup Ready soybeans (resistant to the herbicide glyphosate, commonly called Roundup®)
is consumed. Also, M13 bacteriophage DNA has been shown to survive transiently in the
gastrointestinal tract of mice and is able to enter the bloodstream. However, these studies
demonstrate only the ability of certain DNA molecules to resist degradation by salivary and
gastric enzymes; no studies to date have demonstrated the ability of the DNA molecules to
become stably integrated into the bacterial genome by HGT [50,51].
Some consumers have reported concerns that consumption of bioengineered foods means
that humans will ingest the “foreign” DNA present in transgenes. A DNA sequence of particular
concern is the cauliflower mosaic virus 35S promoter, commonly used to direct expression
of plant transgenes. This promoter is efficient and functional in a variety of organisms, and

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it has been suggested that it might lead to inappropriate over expression of genes in species
into which it is transferred and promote HGT, or recombine with dormant endogenous viruses
present in humans, leading to new infectious viruses. However, almost all genomes of human
endogenous retroviruses contain lethal mutations that prevent replication and production of
viral particles. Also, the cauliflower mosaic virus is present naturally in approximately 10% of
cabbages and cauliflowers, and so is regularly ingested by humans. No adverse consequences
from the consumption of this virus have been reported [50,51].
Food-grade micro-organisms
The use of micro-organisms (Table 1) in food production is accepted when they have a long
history of safe use. However it is not scientifically defined what a long history is and what a
safe use is. Moreover, a recent draft document suggests that only micro-organisms that have a
long and safe history of use in food and that also have a qualified presumption of safety status
(see below) should be applied in the food industry. These strains could be used as recipients for
genetic modification. The use of pathogenic micro-organisms should not be allowed. However,
the history of safe use in food should not be an everlasting guarantee that the strain could
always be applied in food fermentation processes. If new research shows that strains with a
long history of safe use are producing toxic components in levels that may harm human health,
these strains should not be accepted anymore for use in food fermentations. For instance,
certain LAB has an unblemished history of safety in food fermentation, but, as was discovered
later, may produce unfavorable amines under some conditions. Evidently these strains should
not be used in preparation of foods without a profound safety assessment, which, especially in
this case, also investigates the actual concentrations of the harmful compound to which the
consumer will be exposed [50,51].
Host strain Donor strain Gene involved Intended effect Modification
technique
Reference
Lb. bulgaricus Not applicable lacZ limited lactose
fermentation
IS mediated deletion Mollet and Delley,
1990
L. lactis Not applicable ldh and others increased carbon dioxide
production
spontaneous and
induced random
mutagenesis
El Attar et al. 2000
L. lactis Not applicable ldh and others increased acetoin and
diacetyl production
NNG induced random
mutagenesis
Boumerdassi et al.
1997
L. lactis Not applicable aldB increased diacetyl
production
NNG induced random
mutagenesis,
Monnet et al. 2000
L. lactis Not applicable aldB increased diacetyl
production
Spontaneous random
mutagenesis
Goupil et al. 1996
L. lactis Not applicable ribC increased riboflavin
production
induced random
mutagenesis
Burgess et al. 2003
S. thermophilus Not applicable gal operon fermentation of galactose spontaneous random
mutagenesis
Vaughan et al. 2001
L. lactis Not applicable glk, eIIman/glc
no glucose fermenting
capacity
spontaneous random
mutagenesis
Thompson et al.
1985.
L. lactis L. lactis aldB increased diacetyl
production
double crossover
homologous
recombination
Swindell et al. 1996
L. lactis Lb. helveticus pepN, pepX, pepC,
pepI
modulation of proteolytic
system for enhancement
cheese ripening
food grade vector
cloning
Joutsjoki et al. 2002
L. lactis L. delbrueckii pepI, pepL, pepW,
pepG
modulation of proteolytic
system for enhancement
cheese ripening
NICE System Wegmann et al. 1999

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L. lactis Peptostreptococcus
asaccharolyticus
Gdh increased production of
alpha-ketoglutarate
Vector cloning Rijnen et al. 2000
L. lactis lytic phage phi31 phage inducible
promoter
expression of lethal three-
gene restriction cassette
LlaIR+,
Vector cloning Djordjevic et al.1997
L. lactis lytic phage phi31 anti sense phage
RNA
silencing of phage genes Vector cloning Walker and
Klaenhammer, 1998
L. lactis lytic phage anti sense phage
RNA
silencing of phage genes
encoding structural genes
Vector cloning Kim et al. 1992
L. lactis Not applicable pip inactivation of phage
infection protein
double crossover
homologous
recombination
Monteville et al. 1994
L. lactis strains L. lactis strains lacticin encoding
genes
lacticin production Conjugation O’Sullivan et al. 2003
L. lactis strains L. lactis strains lacticin encoding
gene
lacticin production conjugation (plasmid
stacking)
Mills et al. 2002
L. lactis S. thermophilus abiA, abiG abortion of cells upon
phage induction
Vector cloning Tangney and
Fitzgerald, 2002
L. lactis and
others
Pediococcus
acidilactici and
others
lcnC, lcnD lantibiotic production Vector cloning Horn et al. 1999
L. lactis phage lytA, lytH production of lysin and
holin
NICE de Ruyter et al. 1997
L. lactis S. thermophilus Sfi6 EPS gene cluster altered EPS production Vector cloning Stingele et al. 1999
L. lactis S. thermophilus
Sfi39
EPS gene cluster altered EPS production Vector cloning Germond et al. 2001
Lb. gasseri L. lactis folate gene cluster introduction folate
biosynthesis pathway
Vector cloning Wegkamp et al. 2004
S. thermophilus Not applicable pgmA, gal U inactivation of
phosphoglucomutase
double crossover
homologous
recombination and
vector cloning
Levander et al. 2002
L. lactis B. sphaericus ldh, alaD, alr rerouting of pyruvate to
L-alanine
double crossover
homologous
recombination, vector
cloning
Hols et al. 1999
L. lactis L. lactis riboflavin gene
cluster
overexpression riboflavin
Vector cloning Burgess et al. 2003
L. lactis L. lactis folate gene cluster overexpression folate
Vector cloning Sybesma et al.
2003b
L. lactis L. lactis glk, pfnABCD,
pfcBA, genes
lactose-PTS and
tagatose-6P
inactivation of glucose
fermenting system and
introduction of lactose
fermentation
double crossover
homologous
recombination, vector
cloning
Pool et al. 2003
L. lactis L. lactis galA, aga introduction
a-galactosidase activity
food grade vector
cloning
Boucher et al. 2002
Lb. plantarum B. subtilis phyC introduction phytase
activity
Vector cloning Kerovuo and
Tynkkynen, 2000
Lb. plantarum L. amylovorus amyA introduction a-amylase
activity
chromosomal
integration
Fitzsimons et al.
1994
Lb. plantarum B.
stearothermophilus,
C. thermocellum
α-amylase gene,
celA
introduction a-amylase
and cellulase activity
single homologous
recombination
Scheirlinck et al.
1989

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S. thermophilus S. thermophilus glyA increased acetaldehyde
production
Vector cloning Chaves et al. 2002
Lb. fermentum Not applicable ldhD, ldhL increased mannitol
production
double crossover
homologous
recombination
Aarnikunnas et al.
2003
Lb. helveticus Not applicable ldhD production of pure L-(+)
lactic acid
double crossover
homologous
recombination
Kyla-Nikkila et al.
2000
Lb. delbrueckii Not applicable EPS genes altered EPS production Chemically induced
random mutagenesis
Welman et al. 2003
S. thermophilus S. thermophilus
bacteriophage
anti sense phage
RNA, helicase
gene
silencing of phage genes Vector cloning Sturino and
Klaenhammer, 2002
Streptococcus
mutans
Zymomonas mobilis ldh, adh prevention of dental caries double crossover
homologous
recombination, gene
replacement
Hillman, 2002
Lb. delbrueckii Not applicable β-galactosidase
gene
Increased b-galactosidase
activity
EMS or NNG induced
random mutagenesis
Ibrahim and
O'Sullivan, 2000
L. lactis Eimeria tenella, L.
plantarum
M1Pase gene,
MtlD
increased mannitol
production
NICE Wisselink et al. 2005
L. casei ATCC
334
L. casei LC202 dhic increased alpha-keto acid
dehydrogenase activity
Vector cloning Broadbent et al. 2004
L. plantarum Lactobacillus sake promotors and
regulatory genes
increased gene
expression
Vector cloning Mathiesen et al. 2004
L. lactis L. lactis Prt+- and Lac+-
derivatives of L.
lactis MG1363
increased proteolytic and
acidifying activity
conjugation Picon et al. 2005
L. lactis S. simulans Lss production of lysostaphin NICE Mierau et al. 2005
L. lactis P. stipitis XYL1 production of xylitol Vector cloning Nyyssölä et al. 2005
L. lactis Not applicable unknown increased oxidative-stress
tolerance
spontaneous
mutations (natural
selection)
Rochat et al. 2005
L. lactis L. lactis nisRK, nisFEG increased nisn Z
production
Vector cloning Cheigh et al. 2005
L. lactis Z. mobilis, L. lactis pdc, nox increased acetaldehyde
production
NICE Bongers et al. 2005
L. lactis G.
stearothermophilus,
L. Brevis, L. lactis
sgsE, slpA, usp production of excreted
S-layer protein
NICE Novotny et al.2005
L. lactis E. coli gshA, gshB increased glutathion
production
NICE Li et al.2005
L. lactis, L.
paracasei
L. paracasei groEL increased stress tolerance NICE Desmond et al. 2004
L. lactis Not applicable unknown increased L(+)-actate
production
random mutagenesis Bai et al. 2004
S. thermophilus Not applicable deoB, gst, rggC,
and unknown
increased oxidative-stress
tolerance
random insertional
mutagenesis
Fernandez et al.
2004
EMS: ethyl methanesulfonate; NNG: N-methyl-N'-nitro-N-nitrosoguanidine
Table 1: Overview of lactic acid bacteria with controlled or uncontrolled genetic alterations.
Controlled genetic alteration of LAB (Lactic Acid Bacteria)
Relation of a specific aspect to the application of vectors in industrial strain improvement is

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the use of selection markers. The use of antibiotic resistance markers might result in transfer
of antibiotic resistance from one organism to another. As a consequence, the practical value of
antibiotics that are used for human health or in veterinary practice will be severely reduced.
Hence, food-grade resistance markers are preferred. Currently there are many food-grade
selection markers for vector cloning in LAB. For instance, transfer of the α-galactosidase gene
(aga) and a gene coding for a putative transcriptional regulator from the LacI/GalR family
(galR) of Lactococcus raffinolactis ATCC 43920 into L. lactis and Pediococcus acidilactici strains
modifies the sugar fermentation profile from Melibiose negative (Mel(-)) to Melibiose positive
(Mel(+)). A similar food-grade vector is based on complementation of the lactose operon in
L. lactis NZ3600 or L. casei by introduction of lacF, or lacG, respectively, enabling growth
on lactose. An alternative system is based on a suppressor tRNA allowing growth in milk of
a purine auxotrophic strain. A newly developed food-grade marker is characterized by the
requirement of D-alanine in the medium to enable growth of the micro-organisms. An overview
of general strategies for constructing food-grade markers has previously been reported. As
mentioned before, the current use of these food-grade markers if constructed via self-cloning
techniques is restricted to applications with contained use [50,51].
Other targeted modifications of the genetic content of DNA may occur via conjugation
and transduction. These processes are considered natural events. According to the current
legislation, bacteria that are changed by using these transfer systems are not considered as
GMOs (Table 2).
LAB has a long history of use by man for food production and food preservation. LAB is
Gram-positive, non-spore forming bacteria and naturally present in raw food material and
in the human gastro-intestinal tract. The heterogeneous group of LAB includes the rod-
shaped bacteria like lactobacilli, and cocci such as streptococci, lactococci, pediococci and
leuconostocs. LABS are widely used as starter cultures for fermentation in the dairy, meat and
other food industries. Their properties have been used to manufacture products like cheese,
yoghurts, fermented milk products, beverages, sausages, and olives. These food-grade bacteria
can also improve the safety, shelf life, nutritional value, flavor and quality of the product.
Moreover, LAB can be used as cell factories for the production of food additives and aroma
compounds. It is further assumed that LAB may function as probiotics and contribute to the
general health of the consumer upon consumption. The use of probiotics falls currently within
a grey area between food and medicine and many health claims assigned to probiotics are not
yet scientifically proven. Another application - the use of LAB in the production of proteins for
application in health care or for development of new vaccines is more related to Pharma than
to food. In the future it is predicted that knowledge about the interaction between LAB and the
human host will open new avenues for developing LAB which support human health [50-53].
Modification of DNA Directed genetic
alteration
Un-controlled genetic
alteration
Acceptance of contained
use, 90/219/EC
Acceptance of deliberate
release, 2001/18/EC
Spontaneous mutations - + + non-GMO + non-GMO
Induced mutations - + + non-GMO + non-GMO
Mutations via insertion elements - + + non-GMO + non-GMO
Conjugation + - + non-GMO + non-GMO
Transduction + - + non-GMO + non-GMO
Self-cloning + - + non-GMO - GMO
Non-self-cloning + - - GMO - GMO
Table 2: Types of DNA modification methods and the acceptability to be used in food production.

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Conclusions
Claims by a manufacturer that a food or its ingredients, including foods such as raw
agricultural commodities, is not bioengineered should be able to substantiate that the claim
is truthful and not misleading. Available validated testing is the most reliable way to identify
bioengineered foods or food ingredients. For many foods, however, particularly for highly
processed foods such as oils, it may be difficult to differentiate by validated analytical methods
between bioengineered foods and food ingredients and those obtained using traditional breeding
methods. Where tests have been validated and shown to be reliable they may be used. However,
if validated test methods are not available or reliable because of the way foods are produced or
processed, it may be important to document the source of such foods differently. Also, special
handling may be appropriate to maintain segregation of bioengineered and non-bioengineered
foods. In addition, manufacturers should consider appropriate recordkeeping to document
the segregation procedures to ensure that the food’s labeling is not false or misleading. In
some situations, certifications or affidavits from farmers, processors, and others in the food
production and distribution chain may be adequate to document that foods are obtained from
the use of traditional methods. A statement that a food is “free” of bioengineered material may
be difficult to substantiate without testing. Because appropriately validated testing methods
are not currently available for many foods, it is likely that it would be easier to document
handling practices and procedures to substantiate a claim about how the food was processed
than to substantiate a “free” claim.
References
1. Arshad SH, Malmberg E, Krapf K, Hide DW (1991) Clinical and immunological characteristics of Brazil nut allergy. Clin Exp Allergy
21: 373-376.
2. Astwood JD, Leach JN, Fuchs RL (1996) Stability of food allergens to digestion in vitro. Nat Biotechnol 14: 1269-1273.
3. Berberich SA, Ream JE, Jackson TL, Wood R, Stipanovic R, et al. (1996) The composition of insect-protected cottonseed is
equivalent to that of conventional cottonseed. J. Agric. Food Chem 44: 365–371.
4. Betz FS, Hammond BG, Fuchs RL (2000) Safety and advantages of Bacillus thuringiensis-protected plants to control insect pests.
Regul Toxicol Pharmacol 32: 156-173.
5. Carver JD (1999) Dietary nucleotides: Effects on the immune and gastrointestinal systems. Acta Paed 88: 83–88.
6. CDC (2001) Investigation of human health effects associated with potential exposure to genetically modified corn. A Report to the
U.S. Food and Drug Administration from the Centers for Disease Control and Prevention.
7. Conner AJ, Jacobs JM (1999) Genetic engineering of crops as potential source of genetic hazard in the human diet. Mutat Res
443: 223-234.
8. Doerfler W (1991) Patterns of DNA methylation--evolutionary vestiges of foreign DNA inactivation as a host defense mechanism.
A proposal. Biol Chem Hoppe Seyler 372: 557-564.
9. Doerfler W (2000) Foreign DNA in Mammalian Systems. Wiley-VCH, Weinheim.
10. FAO/WHO (2000) Safety aspects of genetically modified foods of plant origin: Report of a Joint FAO/WHO Expert Consultation
on Foods Derived from Biotechnology. Food and Agriculture Organization of the United Nations and World Health Organization.
WHO, Geneva, Switzerland.
11. FAO/WHO (2001) Safety assessments of foods derived from genetically modified microorganisms. Report of a Joint FAO/WHO
Expert Consultation on Foods Derived from Biotechnology. Food and Agriculture Organization of the United Nations and World
Health Organization. WHO, Geneva, Switzerland.
12. FDA (1992) Statement of policy: Foods derived from new plant varieties. Food and Drug Administration. Fed. Reg. 57:22984–
23002.
13. Flavell RB, Dart E, Fuchs RL, Fraley RT (1992) Selectable marker genes: safe for plants? Biotechnology (N Y) 10: 141-144.
14. Gatehouse AM, Ferry N, Raemaekers RJ (2002) The case of the monarch butterfly: a verdict is returned. Trends Genet 18: 249-251.

eBooks
23
15. Goldsbrough AP, Tong Y, Yoder JI (1996) Lc as a non-destructive visual reporter and transposition excision marker gene for
tomato. Plant J. 9:927–933.
16. Hammond BG, Vicini JL, Hartnell GF, Naylor MW, Knight CD, et al. (1996) The feeding value of soybeans fed to rats, chickens,
catfish and dairy cattle is not altered by genetic incorporation of glyphosate tolerance. J Nutr 126: 717-727.
17. Kimber I, Dearman RJ (2001) Can animal models predict food allergenicity? Nutr. Bull 26:127–131.
18. Koprek T, McElroy D, Louwerse J, Williams-Carrier R, Lemaux PG (2000) An efficient method for dispersing Ds elements in the
barley genome as a tool for determining gene function. Plant J 24: 253-263.
19. Kuiper HA, Kleter GA, Noteborn HP, Kok EJ (2001) Assessment of the food safety issues related to genetically modified foods.
Plant J 27: 503-528.
20. Lemaux PG, Frey P (2002) Biotechnology Information.
21. Losey JE, Rayor LS, Carter ME (1999) Transgenic pollen harms monarch larvae. Nature 399: 214.
22. Lozzia GC, Furlanis C, Manachini B, Rigamonti IE (1998) Effects of Bt corn on Rhopalosiphum padi L. (Rhynchota, Aphididae) and
on its predator Chrysoperla carnea Stephen (Neuroptera chrysopidae). Boll. Zool. Agrar. Bachicolt 30:153–164.
23. MacKenzie D (1999) Unpalatable truths. New Sci 162:18–19.
24. Maryanski JH (1995) Food and Drug Administration policy for foods developed by biotechnology. In Genetically Modified Foods:
Safety Issues (K.-H. Engel, G. R. Takeoka, and R. Teranishi, Eds.), pp.12–22. ACS Symposium Series No. 605. American
Chemical Society, Washington, DC.
25. Mayeno AN, Gleich GJ (1994) Eosinophilia-myalgia syndrome and tryptophan production: a cautionary tale. Trends Biotechnol
12: 346-352.
26. Metcalfe DD, Astwood JD, Townsend R, Sampson HA, Taylor SL, Fuchs R, (1996) Assessment of the allergenic potential of foods
derived from genetically engineered crop plants. Crit. Rev. Food Sci. Nutr. 36:165–186.
27. Meyer P, Linn F, Heidmann I, Meyer H, Niedenhof I, et al. (1992) Endogenous and environmental factors influence 35S promoter
methylation of a maize A1 gene construct in transgenic petunia and its colour phenotype. Mol Gen Genet 231: 345-352.
28. Millstone E, Brunner E, Mayer S (1999) Beyond ‘substantial equivalence’. Nature 401: 525-526.
29. American Medical Association Council on Scientific Affairs (2000) CSA Report 10-I-00: Genetically Modified Crops and Foods.
30. Cellini F, Chesson A, Colquhoun I, Constable A, Davies HV, et al. (2004) Unintended effects and their detection in genetically
modified crops. Food Chem Toxicol 42: 1089-1125.
31. Bracket RE for the Food and Drug Administration (2005) Bioengineered foods: Testimony before the Senate Committee on
Agriculture, Nutrition, and Forestry.
32. Food and Drug Administration. Completed consultations on bioengineered foods. Center for Environmental Risk Assessment. GM
crop database.
33. United States Department of Agriculture Economic Research Service. Adoption of genetically engineered crops in the U.S.
34. Byrne P (2010) Labeling of genetically engineered foods. Colorado State University.
35. Food and Drug Administration (2010) Background Document: The VMAC Meeting on Science-Based Issues Associated with Aqua
advantage Salmon.
36. Goodman RE, Vieths S, Sampson HA, Hill D, Ebisawa M, et al. (2008) Allergenicity assessment of genetically modified crops--what
makes sense? Nat Biotechnol 26: 73-81.
37. Herman RA, Ladics GS (2011) Endogenous allergen upregulation: transgenic vs. traditionally bred crops. Food Chem Toxicol 49:
2667-2669.
38. Batista R, Oliveira MM (2009) Facts and fiction of genetically engineered food. Trends Biotechnol 27: 277-286.
39. Ochman H, Lawrence JG, Groisman EA (2000) Lateral gene transfer and the nature of bacterial innovation. Nature 405: 299-304.
40. van den Eede G, Aarts H, Buhk HJ, Corthier G, Flint HJ, et al. (2004) The relevance of gene transfer to the safety of food and feed
derived from genetically modified (GM) plants. Food Chem Toxicol 42: 1127-1156.
41. Tangney M, Fitzgerald GF (2002) AbiA, a lactococcal abortive infection mechanism functioning in Streptococcus thermophilus.
Appl Environ Microbiol 68: 6388-6391.
42. ten Brink B, Damink C, Joosten HM, Huis in ‘t Veld JH (1990) Occurrence and formation of biologically active amines in foods. Int
J Food Microbiol 11: 73-84.

eBooks
24
43. Teuber M, Meile L, Schwarz F (1999) Acquired antibiotic resistance in lactic acid bacteria from food. Antonie Van Leeuwenhoek
76: 115-137.
44. Thompson J, Chassy BM, Egan W (1985) Lactose metabolism in Streptococcus lactis: studies with a mutant lacking glucokinase
and mannose-phosphotransferase activities. J Bacteriol 162: 217-223.
45. Vaughan EE, van den Bogaard PT, Catzeddu P, Kuipers OP, de Vos WM (2001) Activation of silent gal genes in the lac-gal regulon
of Streptococcus thermophilus. J Bacteriol 183: 1184-1194.
46. de Visser JA, Akkermans AD, Hoekstra RF, de Vos WM (2004) Insertion-sequence-mediated mutations isolated during adaptation
to growth and starvation in Lactococcus lactis. Genetics 168: 1145-1157.
47. Von WA, Bruce A (2003) Genetically modified micro-organisms and their potential effects on human health and nutrition. Trends in
Food Science and Technology 14(5-8): 264-276.
48. Walker SA, Klaenhammer TR (1998) Molecular characterization of a phage-inducible middle promoter and its transcriptional
activator from the lactococcal bacteriophage phi31. J Bacteriol 180: 921-931.
49. Wegkamp A, Starrenburg M, de Vos WM, Hugenholtz J, Sybesma W (2004) Transformation of folate-consuming Lactobacillus
gasseri into a folate producer. Appl Environ Microbiol 70: 3146-3148.
50. Wegmann U, Klein JR, Drumm I, Kuipers OP, Henrich B (1999) Introduction of peptidase genes from Lactobacillus delbrueckii
subsp. lactis into Lactococcus lactis and controlled expression. Appl Environ Microbiol 65: 4729-4733.
51. Welman AD, Maddox IS, Archer RH (2003) Screening and selection of exopolysaccharide-producing strains of Lactobacillus
delbrueckii subsp. bulgaricus. J Appl Microbiol 95: 1200-1206.
52. Wisselink HW, Moers AP, Mars AE, Hoefnagel MH, de Vos WM, et al. (2005) Overproduction of heterologous mannitol
1-phosphatase: a key factor for engineering mannitol production by Lactococcus lactis. Appl Environ Microbiol 71: 1507-1514.
53. Wymer P (1998) The application of genetically modified lactic acid bacteria in food products in Europe. Workshop Report, Lactic
Acid Bacterial Industrial Platform (LABIP).

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