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Date:

July 11, 2005

Subject:

Bt is Not Toxic; Are GM Crops Thirsty?; Jared Diamond on TV; Regulatory Regimes for Transgenic Crops

 

Today in AgBioView from www.agbioworld.org : July 11, 2005

* Bt Toxin Not Guilty by Association
* Do Genetically Engineered Crop Varieties Need More Water?
* India: IPR Regime, Public Concern Hamper Growth In Agri-Biotech
* Crops Fail Across Southern Africa
* 'Guns, Germs and Steel'.... Now a PBS Miniseries...Airs Today
* Reply to "Regulatory Regimes for Transgenic Crops"
* ... Regulatory Regimes for Transgenic Crops
--

Bt Toxin Not Guilty by Association

- Ruud A de Maagd, Alejandra Bravo & Neil Crickmore, Nature
Biotechnology 23, 791; July 2005. www.nature.com/nbt ; reproduced in
AgBioView with the permission of the editor.

To the editor: In a Perspective in the September issue (Nat.
Biotechnol. 22, 1105-1109, 2004), Heinemann and Traavik suggested
that horizontal gene transfer from Bt crops (transgenic plants
expressing a cry gene from Bacillus thuringiensis) may pose a food
safety or other environmental hazard because "it is noteworthy that
B. thuringiensis has "a significant history of mammalian
pathogenicity" and is thus not irrelevant to food safety or other
environmental issues." The reference is to a review from us, but
misconstrues our original intent because the original text in our
review actually runs: "Bt does not have a significant history of
mammalian pathogenicity..."1 [emphasis added].

In the April issue, the authors make a correction indicating that
they wrongly cited our reference but persist in their opinion by now
referring to the close relationship between B. thuringiensis,
Bacillus cereus and Bacillus anthracis, of which strains of the
latter two do have significant pathogenicity. In their corrigendum2,
to support their allegation that plants containing a cry gene may
constitute a hazard, they state: "Members of this group are so
closely related that they may be considered members of the same
species, often differing only by the presence or absence of certain
plasmids". Thus, they damn B. thuringiensis, and the use of its
genes, by association with B. cereus, or worse, with B. anthracis,
the causal agent of anthrax.

The authors are correct in noting that B. thuringiensis, B. cereus
and B. anthracis are closely related; they are also correct in noting
that the main differences are often due to the presence or absence of
certain plasmids3. What they fail to note is that in this group of
bacteria, as in many others, host specificity is largely determined
by plasmid-encoded factors. Thus, the distinctive feature of B.
thuringiensis is the production of insecticidal crystal proteins, and
it is only the genes encoding these proteins (which are mostly
plasmid-borne) that are expressed in transgenic plants. Moreover,
these genes are derived from strains of B. thuringiensis that have
been used as safe and environment-friendly sprayable pesticides for
decades, including by many organic farmers.

Furthermore, the focus of Heinemann and Traavik's original
Perspective is on transgenic plants expressing Bt toxin proteins from
the bacterium, not on the bacterium itself. Clearly, the safety of a
single gene product does not inherently reflect that of the complex
organism from which it originates. We are not suggesting that safety
issues should not be considered, but to date there have been many
years of safe Bt toxin use. The overwhelming evidence is that B.
thuringiensis, and transgenic crops expressing cry genes, are not a
threat to mammalian species. The authors of this paper are using a
poor interpretation of good science to imply a poor interpretation
of good science to imply a potentially serious risk.

References
1. de Maagd, R.A., Bravo, A. & Crickmore, N. Trends Genet. 17, 193-199
(2001).
2. Heinemann, J.A. & Traavik, T. Nat. Biotechnol. 23, 488 (2005).
3. Rasko, D.A., Altherr, M.R., Han, C.S. & Ravel, J. FEMS Microbiol.
Rev. 29, 303-329 (2005).

**********************************************

Do Genetically Engineered Crop Varieties Need More Water?

- C Kameswara Rao, Foundation for Biotechnology Awareness and
Education, Bangalore, India; kraovsnl.com
http://www.fbae.org/Channels/Views/why_do_genetically_engineered.htm

The question as to why Genetically Engineered (GE) crops perform
worse than their non-GE counterparts in drought was asked in the
past, and the issue is raised again a few days ago, by the Network of
Concerned Farmers (
http://www.non-gm-farmers.com/news_details.asp?ID=2253).

Soil, temperature and water, when inappropriate, cause what is called
abiotic stress on plants. Farmers know very well that no crop can be
grown without a certain minimal quantity of water, properly
distributed over the crop duration. Crops require more water during
the phases of seed germination, flowering, and fruit and seed
formation. Conventional crop plant varieties are a pampered lot and
cannot withstand water stress (lack of an adequate quantity of water)
for long. They wilt, often beyond the point of recovery, even when
water becomes available subsequently. Such a situation is called
para-wilt, often mistaken for wilt due to pathogens.

Some wild species of plants, such as those growing in the aird
regions, can naturally withstand a physical deficiency of water. Some
other species, growing in saline soils or in mangrove conditions,
tolerate a physiological lack of water, that is, where water is
physically abundant but cannot be utilized by the plants due to high
salt content.

Agricultural scientists have been successful in isolating genes that
provide these species with the ability to withstand a certain degree
of water stress and have developed drought tolerant transgenic crops
containing these genes. We have to distinguish between drought
tolerant and drought resistant crops, both of which would require
much less water than the conventional ones, and no one should expect
even such transgenics to flourish under totally dry conditions.
Drought tolerant plants can withstand only a certain degree of water
stress and there cannot be a GE technology that will make plants grow
without water. The development of drought tolerant transgenics is
still in the early phases and should not be blocked on imaginary and
prompted fears.

Inherent genetic deficiencies, pests, diseases and weeds constitute
biotic stress. High yielding varieties result from a correction of
genetic deficiencies, facilitating a better utilization of inputs
resulting in a higher productivity. Dwarf varieties of rice and wheat
have a reduced vegetative phase, both in volume and time,
facilitating an enhanced availability of inputs for the reproductive
phase resulting in a higher yield. Dwarf stature also prevents
lodging of the crop that causes produce losses. A high yielding
variety requires more water than its counterpart as there is a lot
more of vegetative and reproductive growth, that is greater height of
plants, more branches and leaves, more flowers, fruits and seeds,
called in its entirety, the biomass.

That improved varieties require more water than their counterparts is
not a new issue, only that it was not consciously noticed or
projected as a negative feature earlier. Now it has become handy whip
to beat agricultural biotechnology.

Higher water requirement becomes imperative when we prevent damage
caused by pests and diseases, either by conventional methods or
transgenics. A healthy plant has a greater biomass and demands more
inputs. Conventional methods of cotton pest control were not very
efficient for several reasons. When the most important pest of
cotton, the American bollworm is controlled in Bt varieties, there is
more biomass, which naturally requires more water. Similarly, when we
remove the weeds from a crop field, either manually or through
herbicides, a severe competition for water and other resources is
removed and the resultant increase in biomass needs more water.

Both abiotic and biotic stresses hamper the expression of a crop
variety's full genetic potential. The differences in biomass between
conventional and improved varieties are very obvious when one looks
at, in neighbouring fields, rain-fed and irrigated crop of the same
variety or conventional and improved varieties of the same crop.
Transgenic varieties also mature earlier than their counterparts, as
could be seen with Bt and non-Bt varieties.

The need for additional quantities of water on reducing biotic stress
is naturally more conspicuously manifested in drought conditions,
with added abiotic stress, as there is not only less water available
but also there is more than usual loss of water through evaporation
from the soil and plant surfaces, into the drier environment.

The remedy is in choosing a crop that is right for the abiotic
conditions available in a particular year/crop season. This means
that one cannot grow a choice crop or an improved variety, either all
the years continuously or in all the places. Meteorologists and
agricultural scientists should jointly guide such choices.

**********************************************

India: IPR Regime, Public Concern Hamper Growth In Agri-Biotech

- Ashok B Sharma, Financial Express (India), July 11, 2005

Modern agri-biotechnology, otherwise called transgenic technology in
agriculture, is in a low pace worldwide than compared to bio-pharma.
In India, too, the situation is not different.

Commercialisation of transgenic crops is slated to complete a decade
this year since the commercial sowing of the Flavr Savr tomato in US
in 1995. So far, only 17 transgenic crops have been approved for
planting in various countries. Out of approved transgenic crops, only
four, namely corn, cotton, soybean and canola have a major market
presence. Commercial production of transgenic papaya, squash and
tobacco has been initiated in US. Other transgenic crops such as
chicory, tomatoes, rice, potatoes, flax have been approved for
commercial use in one or more countries, but have not yet been
marketed.

Global area under transgenic crops is seen growing. In 2004, the area
under transgenic crops was reported at 81 million hectares. Yet the
global market size for transgenic crops is only $5 billion as
estimated by International Services for Acquisition of
Agri-biotechnology Application (ISAAA). In contrast, the global
market size for bio-pharma is over $30 billion (2002 estimates).

The only transgenic crop approved in India for commercial cultivation
is Bt cotton. Interestingly, the area under Bt cotton in 2004 was
only 500,000 hectares out of over 9 million hectares under total
cotton crop. The area under Bt cotton is, however, likely to increase
in 2005 as the regulator has approved 13 new Bt cotton hybrids. The
regulator has also approved new areas for commercial cultivation.

A recent study done by BioSpectrum magazine in collaboration with the
Association for Biotech-Led Enterprises (ABLE) has shown that the
market size for bio-agri in India is 6.95% as compared to 75.24% in
case of bio-pharma.

Unlike bio-pharma, agriculture biotechnology has become controversial
particularly in issues relating to health and environmental safety.
This has resulted in slow progress and lower acceptability of
transgenic crops and genetically modified (GM) food.

The intellectual property rights (IPRs) regime is another hurdle in
the path of progress of transgenic technology as identified by the
US-based Pacific Research Institute (PRI). The PRI, which is a
think-tank for market economy, has said that patent-based access and
benefit sharing (ABS) of biological resources is "long-run tax on
biotechnological and pharmaceutical research and development
investment."
The PRI study says that by 2025, the patent-based ABS regime will
reduce biotechnological and pharmaceutical research and development
capital stock by about $144 billion or almost by 27% in 27 select
countries. Alternatively, PRI suggested suggested "contractual
arrangement" for access and benefit sharing of biological resources.

Many developing countries and civil society organisations from the
beginning were against the new IPR regime on life forms. Protection
of traditional knowledge became a necessity when the developed world
and supporters of market economy pushed in the patent regime on life
forms worldwide.
Earlier, the biological resources were freely exchanged between
countries and civilisations. It is of late that the supporters of
market economy have begun realising that new IPR regime on life forms
has become a hurdle.

**********************************************

Crops Fail Across Southern Africa

- BBC NEWS July 09, 2005 http://news.bbc.co.uk/

More than 10 million people need food aid after crop failure in six
southern African countries, the United Nations food agency says. The
World Food Programme says that people are going hungry after erratic
weather, made worse by problems with fertiliser and seeds in some
countries.

Zimbabwe and Malawi are the worst hit countries, the WFP says. It
urged donors to send aid to "avoid widespread hunger from developing
into a humanitarian disaster".

Malawi has experienced its lowest maize harvest since 1992 and will
only cover 37% of average national consumption of 3.4m tonnes of
cereal, the WFP said. In Zimbabwe, the WFP says that four million
people may need aid in the coming year.
Swaziland, Mozambique, Zambia and Lesotho will also need help, it
said on the basis of new crop studies.

All countries are badly hit by the Aids pandemic, which kills those
who would normally be the most productive farmers. Donors say that
Zimbabwe's problems have been made worse by the government's seizure
of white-owned farms. This is strongly denied by the government.

**********************************************

Guns, Germs and Steel.... Now a PBS Miniseries...Airs Today

http://www.pbs.org/gunsgermssteel/

'Guns, Germs and Steel screens across the country on PBS from July 11
for three weeks on Monday nights at 11pm in most cities.'

Based on Jared Diamond's Pulitzer Prize-winning book of the same
name, Guns, Germs and Steel traces humanity's journey over the last
13,000 years - from the dawn of farming at the end of the last Ice
Age to the realities of life in the twenty-first century.

Inspired by a question put to him on the island of Papua New Guinea
more than thirty years ago, Diamond embarks on a world-wide quest to
understand the roots of global inequality.

Why were Europeans the ones to conquer so much of our planet? Why
didn't the Chinese, or the Inca, become masters of the globe instead?
Why did cities first evolve in the Middle East? Why did farming never
emerge in Australia? And why are the tropics now the capital of
global poverty?

As he peeled back the layers of history to uncover fundamental,
environmental factors shaping the destiny of humanity, Diamond found
both his theories and his own endurance tested.

The three one-hour programs were filmed across four continents on
High Definition digital video, and combinied ambitious dramatic
reconstruction with moving documentary footage and computer
animation. They also include contributions from Diamond himself and a
wealth of international historians, archeologists and scientists.

Guns, Germs, and Steel is a thrilling ride through the elemental
forces which have shaped our world -- and which continue to shape our
future.
---------------

First published in the United States by W.W.Norton and Company, on
March 1 1997, Guns, Germs and Steel was initially subtitled 'The
Fates of Human Societies.' Within a few months, this subtitle had
evolved into 'A S

"An epochal work. Diamond has written a summary of human history that
can be accounted, for the time being, as Darwinian in its
authority."-- Thomas M. Disch, The New Leader

"Guns, Germs and Steel lays a foundation for understanding human
history, which makes it fascinating in its own right. Because it
brilliantly describes how chance advantages can lead to early success
in a highly competitive environment, it also offers useful lessons
for the business world and for people interested in why technologies
succeed."--Bill Gates

**********************************************

Reply to "Regulatory Regimes for Transgenic Crops"

- Kent J Bradford, Neal Gutterson, Wayne Parrott, Allen Van
Deynze, & Steven H Strauss, Nature Biotechnology 23, 787 - 789; July
2005. www.nature.com/nbt ; reproduced in AgBioView with the
permission of the editor.

Strauss and colleagues respond:

Wilson et al. claim on the one hand that their report "did not
specifically 'argue for rejection if even a single base pair is
changed,'" while recommending that "transgenic lines containing
genomic alterations at the site of transgene insertion be rejected."
In addition, in their original report, they further state that they
"recommend that both the transgene insertion event (including all
transferred DNA and a large stretch of flanking DNA) and the original
target site be sequenced and compared as the only known way to
definitively determine whether gene sequences have been disrupted."
In the context of their discussion, even a single base pair change is
clearly considered to be a "genomic alteration," so we believe that
we have accurately represented the implications and rationale of
their position.

Regarding the possibility that some genomic changes occur due to
transformation, we never denied that this occurs, and in fact cited
their study as a source for our statement that "unknown mutations and
chromosomal translocations can occur during the transformation and
regeneration process." Where we differ with Wilson et al. is in their
opinion that such mutations will "lead sooner or later to harmful
consequences." There is no documentation of such harmful consequences
in their report for products that have undergone phenotypic screening
for commercial release.

A central point of our Perspective was that a very large number of
genomic and gene differences already exist within crop cultivars, and
even among individual plants within a cultivar, without producing any
harmful consequences (for another striking example, see ref. 1).
Thus, the assumption of the inevitability of harmful consequences
from genomic differences associated with gene transfer ignores the
ubiquity of extensive genome sequence variation within existing food
crops.

Although Wilson et al. agree with us that "analysis of the phenotype
is the one true measure of safety," they nonetheless state that
phenotypic analysis is of "unproven effectiveness" and suggest that
genomic sequence data would be more reliable or effective. Both of
these arguments are flawed. First, phenotypic analysis has been
extremely effective in the development of many thousands of
commercial cultivars in a wide range of crops for several
generations. Second, how Wilson et al. propose to distinguish the
toxicologically silent genomic differences that are abundant in crop
plants from ones that might actually have phenotypic consequences is
addressed neither in their original report nor in their comment.

In his letter, Schubert raises several issues, many of which have
been addressed extensively in published literature. For completeness,
we address these issues here in summary fashion:

Alleged lack of precision in genetic engineering (GE). The lack of
precision due to random gene insertion and genomic alteration is
often raised as a criticism of GE. However, conventional breeding is
based on essentially random induction or assembly of mutations,
followed by selection among a multitude of unpredictable and often
imprecise natural recombinations between genomes. The expression
profile of genes is often changed in ways that are not well
understood, and with multiple phenotypic consequences (that is,
pleiotropy), by inbreeding and wide crosses, as further discussed
below. This lack of 'precision' has not prevented plant breeding from
developing improved crops, as the focus has been primarily on the
resulting phenotypes, not on their genomic basis. Similarly,
ancillary genomic changes accompanying GE may occur, but are
irrelevant so long as the expected and desired phenotype is produced
without unacceptable side effects.

Basic research versus cultivar development. Schubert cites extensive
"unintended effects," but many of these result from failing to
distinguish between the use of transgenes in basic research and the
development of improved cultivars using GE. Unexpected changes in
phenotypes, usually due to overexpression or knockouts, are a routine
part of basic research using GE. However, these events are not
subjected to the phenotypic, biochemical and often molecular
selection demanded in breeding of competitive crop varieties.
Breeders, whether working with conventional methods or transgenes,
conduct years of intensive laboratory, greenhouse and field screens
so phenotypically abnormal, unstable or undesirable genotypes or
events are discarded.

Prevalence of mutagenized cultivars. Schubert states "mutagenesis was
used in the United States during the middle part of the past century,
but food crops made by this technique now constitute less than a few
percent of US production, with sunflowers being the major
representative," citing ref. 2. This is a rather disingenuous summary
of the cited paper, which documents the extensive use and enormous
economic impact of the more than 2,275 varieties of 175 species that
have been derived either as direct mutants or from their progenies.
Many currently popular varieties of numerous crops contain
mutagenized progenitors in their pedigrees. The widespread production
and consumption of mutation-derived varieties without ill effect over
the past 50 years is evidence that these do not need to be regulated
differently from varieties developed via other methods.

Wide crosses and ploidy manipulation. Schubert goes on in his letter
to argue that conventional breeding is inherently safer than GE,
stating that "in wide crosses and other forms of ploidy manipulation,
there are clearly changes in gene dosage, and proteins unique to only
one parent can be produced in the hybrid, but there is no a priori
reason to assume that mutations are going to occur simply because
there is a change in chromosome or gene number." Rather than relying
on a priori assumptions, a large body of evidence indicates that
complex and as yet poorly understood genetic changes often accompany
wide crosses and ploidy manipulation, including gain and loss of DNA,
gene silencing, translocations, epigenetic modifications and
mobilization of transposable elements (e.g., refs. 3, 4, 5, 6).
Schubert's statement that "only GE and mutagenesis introduce large
numbers of mutations" is grossly incorrect. In addition,
introgression of genes via wide crosses most often occurs via
recombination and substitution of chromosomal segments, not via
increases in ploidy, as Schubert claims.

Dangerous nature of genetic changes? Schubert writes that "Strauss
and colleagues correctly state that plants normally contain the same
Agrobacterium tumefaciens and viral DNA sequences that are used to
create GE transfection constructs, but fail to point out that with GE
these pieces of DNA are part of a cassette of genes for drug
resistance along with strong constitutive viral promoters...which are
used to express foreign proteins at high levels in all parts of the
plant--hardly a natural event." This argument has several problems.
First, strong promoters are not restricted to viral DNA; plants also
naturally contain many strong, near-constitutive promoters (e.g.,
ref. 7), and some of these are now used to aid plant transformation
(e.g., refs 8,9).

Second, the viral promoters/enhancers Schubert is concerned about act
over very limited distances on a genomic scale, and thus have very
limited potential to cause random increases of gene expression. The
fourfold repeated cauliflower mosaic virus enhancer element (the
source of its constitutive promoter activity) influences gene
expression predominantly over 5 kb10, or about the size of a single
genomic locus in plants. Third, the use of tissue-specific,
plant-derived promoters, rather than constitutive promoters, is
becoming increasingly common in GE programs (e.g., refs 11,12).
Fourth, those transgenic crops that express antibiotic resistance
genes (not all transgenic crops do) express only those genes whose
expression is already widespread in bacteria found in the human gut
(e.g., refs 13, 14, 15). Finally, with respect to drug resistance
marker genes generally, an in-depth review recently concluded "that
there are no objective scientific grounds to believe that bacterial
AR [antibiotic resistance] genes will migrate from GM plants to
bacteria to create new clinical problems16."

Retrotransposons. Schubert claims that our statement that
"retrotransposons continuously insert themselves between genes" is
incorrect because these high copy number elements are
transpositionally inactive in normal modern food plants. The latter
statement is not supported by experimental results. Expressed
sequence tag databases reveal that retrotransposon RNA is present in
plants17, 18, 19, from which it can only be inferred that their
expression continues. The rate of transposition is likely to be
highly variable depending on species, developmental stage and
inducers, such as environmental and genomic stress. Common non-GE
procedures such as tissue culture, which is used routinely for
dihaploid production and propagation, are known to substantially
increase the rate of transposition (e.g., ref. 20), and many tissue
culture?derived, non-GE varieties have been in the food supply for
some time.

Screening for unexpected molecules. The high diversity of
"nonessential small molecules that provide adaptive benefits under
conditions of environmental or predator-based stress" that Schubert
refers to are also produced in complex and unpredictable ways during
normal crop management, shipping, storage, processing and food
preparation. Cheeses, plant-derived beverages and many other
processed foods are known to contain vast numbers of biochemicals of
diverse types (e.g., refs. 21), the great majority of which have
never been tested for safety. Should all the molecules produced by
each new type of cheese be subject to detailed toxicological
assessments? This also underlines the general, rather than specific,
basis of human adaptation to diverse plant chemistries. Human
digestive systems routinely deal with vast numbers of natural
chemicals present at low concentrations in food, many of which can be
shown to be mutagenic at high concentrations22.

The nucleic acid or proteomic tests of large numbers of gene
expression products that were proposed by Schubert are extremely
sensitive and extremely expensive. They may detect hundreds or even
thousands of changes in a novel variety, whether conventionally bred
or produced using GE, if compared with their progenitors under a full
range of growth environments, stresses and developmental stages. How
would such data be interpreted with respect to risk? Simply obtaining
more data via mandated mass spectrometry, microarray evaluations or
the like, without a means to evaluate them with respect to
benefit/risk of whole foods, does not add to knowledge and safety but
to chaos and controversy.

Schubert backs up his argument by noting that Kuiper et al.23 called
for metabolic profiling of each transgenic event. However, coauthors
of that paper now agree24 that "further research is required to
validate profiling methodologies...The safety assessment of
[genetically modified] GM crops should focus primarily on the
intended novel traits (target gene(s) and product(s)). Unintended
effects occur in both GM and non-GM crops; however, GM crops are
better characterised. It may be suggested that the two should be
treated the same in safety assessments, bearing in mind that safety
assessments are not required for non-GM crops. Profiling techniques
should not at present be an official requirement24."

Finally, because random mutations and alterations in gene expression
occur widely in all plants during breeding, if perturbations of
biosynthetic pathways could readily give rise to important toxins
from commonly grown crops their effects should already be widely
observed. Experience indicates, however, that phenotypes and
metabolic pathways tend to be highly buffered from the effects of
mutations. This is likely to be the reason that most loss-of-function
mutations show only minor, if any, phenotypic changes. For example,
in a screen for insertional inactivation in Arabidopsis thaliana,
only 3% of the T-DNA insertions among a population of 55,000 events
showed a visible phenotype25. This buffering appears to be due to the
immense number of interactions and feedback mechanisms in higher
organisms26, which can occur at the levels of gene expression,
enzymatic pathways, cellular processing and multicellular development.

Unintended changes in plant composition. To support his contention
that unintended consequences can arise from GE, Schubert cites one
study that found higher lignin levels in transgenic Bt maize.
However, those results were not reproduced in a more extensive
study27. Numerous studies document the equivalent performance of
animals fed silage from Bt and non-Bt corn28, 29, 30 (reviewed in
ref. 31), which would not be expected were their lignin compositions
substantially altered.

Likewise, Schubert cites the claim that isoflavone levels are altered
in transgenic soybeans. This claim has been roundly criticized
because it did not compare soybeans of the same genetic background or
grown in the same environment, two factors that are known to have a
large effect on isoflavone content (see
http://www.soybean.com/gmsoyst1.htm). The example of isoflavone
variability in soybean also illustrates the fallacy behind testing
for metabolites; merely finding a difference in the amounts of
metabolites is biologically irrelevant without additional information
on the beneficial versus deleterious effects of specific metabolites
in whole plants and on the range of metabolite levels that can occur
within different genotypes grown under a wide range of environmental
conditions24.

Value of mutagenicity tests. Schubert suggests use of the Ames test,
apparently to examine whether "unexpected changes in small-molecule
metabolism" are of mutagenic significance. However, it is widely
known that this high-dose test gives a greatly inflated rate of false
discovery of nontoxic minor compounds in food (e.g., approximately
half of the compounds in coffee do not pass this test22). The results
of these tests are also known to be very poor predictors of the
potential for mammalian carcinogenicity32. Compounds that are harmful
at the high concentrations used in such tests may even be beneficial
to health at low concentrations. Given the hundreds of metabolites
that may be altered via conventional or GM breeding (not to mention
by environmental conditions, or the presence of pathogens or
insects), it is exceedingly unlikely that screening them via the Ames
test would contribute to the goal of producing more healthful foods.

Our article attempted both to put recombinant DNA modification in a
genomic context with respect to traditional breeding methods and the
diversity of wild progenitors and to propose a regulatory framework
where the benefits from use of gene transfer approaches are not lost
amidst excessive attention to collateral genomic changes. Unintended
genomic changes can be significant for all forms of breeding,
including gene transfer. Yet the preponderance of scientific
research, and experience from plant breeding and applied
biotechnology, suggests that the effects of these genomic changes on
food safety are modest and manageable by paying attention to plant
phenotypes. The technical and ethical challenge is to distinguish
important risks from trivial ones so the many tangible benefits that
can be provided by GE are not stifled by burdensome regulatory
requirements that do not enhance safety of the food supply.

References
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2. Ahloowalia, B.S. et al. Euphytica 135, 187-204 (2004).
3. Liu, B. & Wendel, J.F. Genome 43, 874?880 (2000).
4. Levy, A.A. & Feldman, M. Biol. J. Linn. Soc. 82, 607-613 (2004).
5. Pires, J.C. et al. Biol. J. Linn. Soc. 82, 675-688 (2004).
6. Madlung, A. et al. Plant J. 41, 221-230 (2005).
7. Wang, J. & Oard, J.H. Plant Cell Rep. 22, 129-134 (2003).
8. Zhang W., McElroy, D.R. & Wu, R. Plant Cell 3, 1155?1165 (1991).
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11. Kasuga, M. et al. Nat. Biotechnol. 17, 287-291 (1999).
12. Garg, A.K. et al. Proc. Natl. Acad. Sci. USA. 99, 15898-15903
(2002).
13. Berche, P. Méd. Thér. 4, 709?719 (1998).
14. Calva J.J., Sifuentes Osornio J. & Ceron, C. Antimicrob. Agents
Chemother. 40, 1699 (1996).
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16. Bennett, P.M. et al. J. Antimicrob. Chemother. 53, 418-431
(2004).
17. Echenique, V. et al. Theor. Appl. Genet. 104, 840?844 (2002). |
18. Neumann P., Pozarkova, D. & Macas, J. Plant Mol. Biol. 53,
399?410 (2003).
19. Kuhl, J.C. et al. Plant Cell 16, 114?125 (2004).
20. Hirochika, H. et al. Proc. Natl. Acad. Sci. USA 93, 7783-7788
(1996).
21. Sablé, S. & Cottenceau, G. J. Agric. Food Chem. 47, 4825-4836
(1999).
22. Ames, B.N. & Gold, L.S. FASEB J. 11, 1041-1052 (1997).
23. Kuiper, H.A. et al. Plant J. 27, 503?528 (2001).
24. Cellini, F. et al. Food Chem Toxicol. 42, 1089-1125 (2004).
25. Chaudhury, A. et al. Plant Cell 11, 1817-1825 (1999).
26. Daenicke, R., Aulrich, K. & Flachowsky, G. Mais, September,
135-137 (1999).
27. Jung, H.G. & Sheaffer, C.C. Crop Sci. 44, 1781-1789 (2004).
28. Siegel, M.L & Bergman, A. Proc. Natl. Acad. Sci. USA 99, 10528
(2002).
29. Russell, J.R. et al. in 2000 Beef Research Report--Iowa State
University, pp. 56-61.
30. Barriere, Y. et al. J. Dairy Sci. 84, 1863-1871 (2001).
31. Van Deynze, A.E. et al. Crop Biotechnology: Feeds for Livestock.
(University of California Division of Agriculture and Natural
Resources, Oakland, CA, 2004).
http://anrcatalog.ucdavis.edu/pdf/8145.pdf. Supplemental References:
http://sbc.ucdavis.edu/Publications/8145_Supplement.htm
32. Bethel, A. et al. Environ. Mol. Mutagen. 29, 312?322 (1997).

**********

Regulatory Regimes for Transgenic Crops

- Allison Wilson, Jonathan Latham & Ricarda Steinbrecher Nature
Biotechnology 23, 785; July 2005. www.nature.com/nbt ; reproduced in
AgBioView with the permission of the editor.

To the editor: In presenting their justifications for reducing the
regulatory burden on transgenic food crops (Nat. Biotechnol. 23,
439?444, 2005), we feel that Strauss and colleagues significantly
misrepresent the implications and rationale of our report Genome
Scrambling-Myth or Reality? Transformation-Induced Mutations in
Transgenic Crop Plants1. Unlike their characterization of our work,
we did not specifically "argue for rejection if even a single base
pair is changed." In full, our relevant recommendations were that
"transgenic lines containing genomic alterations at the site of
transgene insertions be rejected" and that "the insertion of
superfluous DNA be considered unacceptable."

Leaving aside the fact that a single base pair change may result in
serious phenotypic consequences, these recommendations are best
viewed in context. As documented in the report, thorough analysis
reveals that all particle bombardment transgene insertion events
include extensive rearrangements or loss of host DNA as well as
insertion of superfluous DNA. Furthermore, a large fraction of even
apparently simple Agrobacterium tumefaciens-mediated transgene
insertion events also result in large-scale host DNA rearrangement or
deletion and superfluous DNA insertion2. For example, loss of 76 kbp
of host DNA3 and duplication/translocation of up to 40 kbp of host
DNA have been reported at T-DNA insertion sites4.

Widespread use of transgenic crops carrying insertion-site mutations
of this magnitude will, in our opinion, lead sooner or later to
harmful consequences. Nevertheless, detailed inspection has shown
that mutations such as these would almost certainly pass unnoticed
through both the molecular and phenotypic characterization stages of
the regulatory systems of both the European Union and the United
States5, 6, 7, 8.

We do agree with Strauss and colleagues that analysis of the
phenotype is the one true measure of safety. However, rigorous
assessment only at the phenotypic level is time consuming, expensive
and, more importantly, of unproven effectiveness9. In this context,
our recommendations for the detection and elimination of
transformation-induced mutations from commercial crop plants are
conceived as a straightforward and effective way to reduce the
probability of unexpected deleterious phenotypes arising in
transgenic crop plants and of protecting consumers and others from an
unnecessary risk.

References - see original paper

********

Regulatory Regimes for Transgenic Crops

- David Schubert, Nature Biotechnology 23, 785 - 787; July 2005.
www.nature.com/nbt ; reproduced in AgBioView with the permission of
the editor.

To the editor: In the April issue (Nat. Biotechnol. 23, 439-444,
2005), Strauss and colleagues argue that the methods used to produce
food crops should not be the focus of regulatory oversight, only the
phenotypic traits of the resultant plants as defined in terms of
standard agricultural practice. They propose that any risk and safety
assessments of crops produced by genetic engineering (GE) should be
based only upon the nature of the introduced genes. They also claim
that transgenic crops face a "daunting" array of regulatory
requirements.

However, safety testing requirements in the United States are largely
voluntary and in my view inadequate (for a review of regulations from
my perspective, see ref. 1). Safety concerns related to the GE
process itself as well as its unintended consequences are set aside
by Strauss and colleagues as irrelevant, for they claim that the
products of genetic events that occur naturally and with standard
plant breeding techniques are fundamentally the same as those that
occur with GE. Are these arguments a valid reflection of what is
known about the precision and consequences of the GE process compared
with naturally occurring genomic variation?

The basic assumption underlying the concept of a one-to-one
relationship between the transgene and the resultant phenotype is
that the GE process is relatively precise. However, none of the
current transgene insertion techniques permits control over the
location of the insertion site or the number and orientation of the
genes inserted. Indeed, over one-third of all Agrobacterium
tumefaciens-mediated insertion events disrupt functional DNA2, 3.
These and related transformation and cell culture-induced changes in
chromosomal structure have been recently documented in great detail4.
For example, translocations of up to 40 kb5, scrambling of transgene
and genomic DNA6, large-scale deletions of over a dozen genes7 and
frequent random insertions of plasmid DNA8 can all be caused by the
procedures used to make transgenic plants. In fact, the most commonly
used transformation procedure is sometimes itself used as a mutagen9
and can activate dormant retrotransposons that are mutagenic10.
Moreover, mutations linked to the transgene insertion site cannot be
removed by additional breeding as long as there is selection for the
transgene itself. Collectively, these data indicate that the GE
process itself is highly mutagenic.

Some modern breeding technologies introduce new traits into plants
via chemical or radiation mutagenesis or by wide cross-hybridizations
that overcome natural species barriers. Mutagenesis was used in the
United States during the middle part of the past century, but food
crops made by this technique now constitute less than a few percent
of US production, with sunflowers being the major representative11.
However, plants produced by wide crosses, such as those between
quackgrass and bread wheat to yield a widely planted grain that has
all of the chromosomes of wheat and an extra half genome of the
quackgrass, although unique, are fundamentally different from those
produced by either mutagenesis or GE. In wide crosses and other forms
of ploidy manipulation, there are clearly changes in gene dosage, and
proteins unique to only one parent can be produced in the hybrid, but
there is no a priori reason to assume that mutations are going to
occur simply because there is a change in chromosome or gene number.
Although the extent and suddenness of all of these modern breeding
technologies are unlike anything known to occur during the course of
evolution or with traditional breeding, only GE and mutagenesis
introduce large numbers of mutations. Any new cultivars derived by
the latter two methods should be subjected to similar regulatory
requirements.

Strauss and colleagues correctly state that plants normally contain
the same A. tumefaciens and viral DNA sequences that are used to
create GE transfection constructs, but fail to point out that with GE
these pieces of DNA are part of a cassette of genes for drug
resistance, commonly along with strong constitutive viral promoters
(e.g., cauliflower mosaic virus promoter), which are used to express
foreign proteins at high levels in all parts of the plant-hardly a
natural event. They incorrectly imply that changes in ploidy, gene
copy number, recombination and high genomic densities of transposable
elements in normal plants continually lead to mutations and changes
in gene expression similar to those caused by GE.

Ploidy is notoriously unstable in plants, but changes involve moving
around large blocks of intact genes while maintaining their regulated
expression pattern. It should also be remembered that recombination
is not the same as random mutagenesis, for there has been tremendous
selective pressure for alleles to express functionally similar
proteins. The statement that "retrotransposons continuously insert
themselves between genes" is incorrect, for these high-copy number
elements are very rarely transpositionally active in normal modern
food plants12, have evolved and rearranged in the distant past13, but
can be activated by tissue culture or by mutagenesis10. In fact,
their discovery by Barbara McClintock was facilitated by the use of
mutagenized corn12.

In contrast to Strauss and colleagues' proposal that regulatory
efforts should focus on the expression of the transgene, I believe
that the potential negative impact on nutritional content or increase
in dangerous metabolites are the major hazards associated with highly
mutagenic plant transformation techniques. Although it is widely
recognized that the breeding of some crops can produce varieties with
harmful characteristics, millennia of experience have identified
these crops, and breeders test new cultivars for known harmful
compounds, such as alkaloids in potatoes14, 15. In contrast,
unintended consequences arising from the random and extensive
mutagenesis caused by GE techniques opens far wider possibilities of
producing novel, toxic or mutagenic compounds in all sorts of crops.
Unlike animals, plants accumulate thousands of nonessential small
molecules that provide adaptive benefits under conditions of
environmental or predator-based stress16. Estimates are that they can
make between 90,000 and 200,000 phytochemicals with up to 5,000 in
one species17. These compounds are frequently made by enzymes with
low substrate specificity18 in which mutations can readily alter
substrate preference19, 20.

There are many examples of unpredictable alterations in
small-molecule metabolism in transgenic organisms. In a yeast strain
genetically engineered to increase glucose metabolism, the
transformation event caused the unintended accumulation of a highly
toxic and mutagenic 2-oxoaldehyde called methylglyoxal21. In a study
of just 88 metabolites in three groups of potatoes transformed with
genes for bacterial and yeast enzymes that alter sucrose metabolism.
Roessner et al.17 found that the amounts of the majority of these
metabolites were significantly altered relative to controls. In
addition, nine of the metabolites detected in these transgenic
potatoes were not detected in conventional potatoes. Given the
enormous pool of plant metabolites, the observation that 10% of those
assayed are new in one set of transfections strongly suggests that
undesirable or harmful metabolites may be produced and accumulate22.
Contrary to the suggestions of Strauss and colleagues, Kuiper et
al.23 strongly recommend that each transformation event should be
assayed for these types of unintended events by metabolic profiling.

A well-documented horticultural example of unintended effects is the
alteration in the shikimic acid pathway in Bacillus thuringiensis
(Bt) toxin corn hybrids derived from Monsanto's MON810 and Syngenta's
Bt11 plants as well as glyphosate-tolerant soybeans. Stem tissue of
both groups of plants has elevated levels of lignin, an abundant
nondigestible woody component that makes the plants less nutritious
for animal feed24, 25. Components of this same biochemical pathway
also produce both flavonoids and isoflavonoids that have a high
nutritional value, and rotenone, a plant-produced insecticide that
has been associated with Parkinson disease26. Isoflavonoids are
abundant in legumes like soy beans, and rotenone is synthesized
directly from isoflavones in many legume species27. Because of the
promiscuity of many plant enzymes and the large and varied substrate
pools of phytochemical intermediates, it is impossible to predict the
products of enzymes or regulatory genes mutated during the
transformation event22. Although I am not aware of any testing of GE
soybeans for rotenone, it has been shown that glyphosate-tolerant
soybeans sprayed with glyphosate have a reduced flavonoid content28.

The safety testing of GE crops need not be as extensive as that done
with drugs, food additives or cosmetics. Many suggestions have been
put forward (e.g., see refs. 1,4,23,29) including those by the World
Health Organization30. I believe that the most important safety tests
include metabolic profiling to detect unexpected changes in
small-molecule metabolism23 and the Ames test to detect mutagens31.
Molecular analysis of the gene insertion sites and
transformation-induced mutations4 should also be performed along with
both multigenerational feeding trials in rodents to assay for
teratogenic effects and developmental problems, and allergenicity
testing performed according to a single rigorous protocol30. The
animal studies are of particular importance for crops engineered to
produce precursors to highly biologically active compounds, such as
vitamin A and retinoic acid, molecules that can act as teratogens at
high doses32.

In summary, Strauss and colleagues state that there is a low risk
from the consumption of transgenic plants "where no novel biochemical
or enzymatic functions are imparted." The question is, of course, how
can one know if a novel and potentially harmful molecule has been
created unless the testing has been done? How can one predict the
risk in the absence of an assay? Because of the high mutagenicity of
the transformation procedures used in GE, the assumptions made by
Strauss and colleagues and by the US Food and Drug Administration33
about the precision and specificity of plant genetic engineering are
incorrect. Nonetheless, it appears that the position of Strauss and
colleagues and the agbiotech industry, as well as the current US
regulatory framework for the labeling and safety testing of
transgenic food crops, is to maintain the status quo and hope for the
best.

The problem is that there are no mandatory safety testing
requirements for unintended effects and that it may take many years
before any symptoms of a disease arising from a transgenic product to
appear. In the absence of strong epidemiology or clinical trials, any
health problem associated with an illness caused by a transgenic food
is going to be very difficult, if not impossible, to detect unless it
is a disease that is unique or normally very rare. Therefore,
although GE may enhance world health and food crop production, its
full potential may remain unfulfilled unless rigorous prerelease
safety testing can provide some assurance to consumers that the
products of this new technology are safe to eat.

For references - see original paper

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