ADVANCES IN LIFE

SCIENCES

Editors
Kajeshwar P. Sinha
Centre of Advanced Study in Botany,
Banaras Hindu University, Varanasi, India

Naveen K. Sharma
Department of Botany, Postgraduate College
Ghazipur, India

Ashwani K. Kai
Centre of Advanced Study in Botany,
Banaras Hindu University, Varanasi, India

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 15
Problems, Pitfalls and Advantages of
Transgenic Crops
PANKAJ KUMAR1 AND ASHWANI K. RA~"

IDeparlment of Botany, UP Autonomous College, Varanasi-221 001, India

2Deparlment of Botany, Banaras Hindu University, Varanasi-221 005, India
*e-mai/:akrai.bhu@gmai/.com

ABSTRACT
I. INTRODUCTION
2. TRANSGENICS AND BIODIVERSITY
3. EFFECT ON NON-TARGET ORGANISMS
4. BIOSAFETY OF TRANSGENIC FOODS
5. BRINGING WASTELAND UNDER CULTIVATION
6. TRANSGENICS AND GENE FLOW
7. ENVIRONMENTAL RISKS ASSOCIATED WITH EVOLUTION OF RESISTANCE
7.1. Physiology and Genetics of Resistance
7.2. Why are the Predictions Wrong?
7.3. Fitness of Resistant Individuals
8. REDUCING PESTICIDES USAGES AND POLLUTIONS
8.1. Monitoring, Detections and Detoxifications of Environmental Pollution
9. CONCLUDING REMARKS
References
338 Advances in Life Sciences

Keywords: Biodiversity, environmental pollution, gene flow, phytoremediation, transgenics,

unintended effects.

1. INTRODUCTION

The advent of recombinant DNA technique enabled the development of the first transgenic crop
(also known as genetically modified crops; GM) in 1987. It involved transfer and expression of
insecticidal crystal protein (cry) gene of Bacillus thuringiensis (Bt) in tobacco (Hilder et al.,
1987; Vaeck et al., 1987). Since then, more than 30 plant species have been transformed for
traits like enrichment of nutrition, production of edible vaccines, delayed ripening processes and
resistance to insects, diseases and herbicides. The growing success of transgenic crops is evident
by increase in global area under transgenics plantation from 1.7 million hectare in 1996 to 90
million hectare in 2005 (Bhalla 2006).
Expansion of transgenic crops is likely to shift market from current generation of 'input'
agronomic traits to the next generation of 'output' quality traits (Ghosh 2001). For example,
development of Golden Rice- 2 (paine et al., 2005) and plants with elevated vitamin E content
(Ajjawi & Shintani 2004). However, besides an array of promises offered transgenics as well as
their products often present new ecological and health concerns. Amidst loud and contentious
voices against the transgenics, in recent times, a set of straightforward, scientifically based
concern has evolved dictating for a cautious approach in creating the best choices for future
agriculture. This article is an attempt to critically analyze the projected conjectural risks of
transgenics in light of available scientific knowledge.

2. TRANSGBNICS AND BIODIVERSITY

Increase in genetic uniformity and high invasiveness are the main concerns regarding the effect
of transgenic crops on biodiversity. Gepts & Pappa (2003) held GM crops responsible for decline
of genetic diversity within a crop species. In contrast; the subsidiary body of Scientific Technical
and Technological Advice (SBSTTA) of the Convention on Biological Diversity (CBD) ranked
habitat loss as the greatest threat to the global biodiversity. If current trends continue, the world
population is expected to exceed 10 billion by 2050. This will require doubling the food
production either from the same amount of arable land (Tilman et al., 2002) or by increasing
 Problems, Pitfalls and Advantages of Transgenic Crops 339

the area under cultivation. Conversion of natural ecosystems to agriculture will accelerate habitat
loss and have negative impact on biodiversity (Jenkin 2003). The plant breeding and genetic
engineering techniques that help to produce more food from a given piece of land, therefore, .
limiting the expansion of cultivation to new areas are urgently required (Hegde & Ellstrad
2002). In this strategy, the role of transgenic plants appears very promising. For soybean and
cotton, it has been reported that genetic uniformity actually decreased after introduction of
transgenic cultivars (Sneller 2003; Bowman et al., 2003).
Role of tillage practice in accelerating biodiversity loss has long been recognized, which
results in degradation and erosion of soils. With the introduction of effective herbicides
dependence on tillage reduces, therefore; use of transgenics tolerant to herbicides provides new
tool for controlling weeds, and adoption of conservation tillage practice and no-till practices
(Ammann 2005). Regarding high invasiveness of the transgenics, Crawley et at. (1993) reported
that transgenic oilseed rape (Brassica napus sub sp. oleifera) were less persistent and invasive
than their conventional counterpart.

3. BFFBCT ON NON-TARGBT ORGANISMS

An argument quoted very much against the widespread use of transgenics is that of their effect
on non-target organisms. For example, Hilbeck (1998) reported positive impact of Bt toxin on
lacewing larvae however; the very result was later on disproved by Romeis et at. (2004). Similarly,
Losey et at. (1999) reported that transgenic pollens harm Monarch butterfly larvae; a non-target
organism. Though, subsequent field experiments demonstrated no significant impact of the Bt
protein on Monarch larvae (Jesse & Obrycki 2000; Ortman 2001). In conventional methods of
pesticide use, besides the target pests many non-target species are also affected, resulting in low
population of non-target organisms (Candolfi et al., 2004). With the expression of insecticidal
proteins in Bt crops, there will be limited activity range with little or no activity against non-
target organisms (Ammann 2005). Moreover, expression of toxin within plant tissues minimizes
the exposure of animals that do not feed on the crop plants. Lozzia et at. (1998) while studying
the effect of GM crops on non-target organisms found that indices of species diversity and
community structure are little affected. In a comparative study, Reed et at. (2001) reported
larger population of many generalist predators in Bt potato fields compared to that of conventional
potato fields treated with appropriate broad spectrum insecticides.

4. BIOSAFBTY OF TRANSGBNIC FOODS

GM crop derived foods due to their unintended effects are often perceived disparingly by
consumers. Unintended effects are changes in chemical composition of the GM plant compared
to that of non-GM plant grown under similar conditions. These changes might affect the potential
agronomic performance but mayor may not pose safety threats to human health and environment.
In unintended effects; effects that could be expected on the basis of our current knowledge
about plant metabolism, gene-gene interaction and general plant biology are known as predictable
unintended effects whereas, those that cannot be anticipated by our current knowledge are
unpredictable effects. Allergenicity, toxicity and possibility of transferring antibiotic resistance
340 Advances in Life Sciences

are the major health concerns associated with transgenic foods (Lack 2002). GM foods may
cause allergy, when an allergen is transformed from allergenic host crop to a non-allergenic
target crop or due to the formation of a new component by insert characteristics. The allergy'
resulting from the insertion of allergenic sequence could be diagnosed by doing 19B-binding
studies, and taking into account physico-chemical characteristic of proteins and thereafter referring
to know allergen databases. However, in former case, it is difficult to assess the risk associated
with the transgenic foods. Furthermore, genetic modification could inadvertently enhance natural
plant toxins by switching on a gene that has the desired effect and capacity to pump out a
poison (Uzogara 2000).
Inserted foreign genes may alter nutritional value of foods in unpredictable way by decreasing
the level of some nutrients while increasing the others. This results in a difference between
traditional strain and its GM-counterpart. However, little is known the way these altered nutrients
are involved in complex regulation of gene expression (Young & Lewis 1995). It is a general
belief that changes in food and diet through biotechnology occur at a pace far greater than the
scientists' ability to predict the significance of such changes. Besides these concerns, consumers
also have a genuine fear of the 'unknown'. Since consuming GM foods that had never before
existed in nature and that too, without prior testing and labeling would reduce consumers to
mere guinea pigs in a colossal biological experiment. However, the proponents of GM food
maintain that potential benefits of GM food are almost limitless and can solve many problems
of world agriculture, health and ecology. They argue that medical biotechnology, which accounts
for the majority of the products of genetic engineering encounters the least controversy, while
food biotechnology often faces criticism that are entirely speculative.
The 'substantial equivalence' concept for the redressal of perceived fear from GM foods
embodies. that: If the rDNA product is substantially equivalent to an existing product, safety
assessment of the rDNA product should be made that the novel food is essentially the same as
that found in nature except for the novel trait. The concept is not new as the same principle is
used in assessing the safety of new hybrid varieties produced by non-biotechnological means.
Decision tree approach can also be applied to assess potential allergenicity of the GM foods
before bringing to the market.
There is a growing demand of 'organic foods' primarily by the consumer's perception of
their quality and safety. However, the 'organic' label is not a health claim, but a process claim.
It denotes products that have been produced in accordance with certain standards during food
production, handling, processing and marketing stages and certified by a duly constituted
certification body or authority. It should not necessarily be interpreted to mean that the foods
produced are healthy, safe or all natural (Ramesh et al., 2005). Avery (1998) reported that
consuming organic foods increases exposure to microbiological contaminants however, there is
no evidence to support such claims (Burros 1999; Pell 1997). Likewise, the report that several
self-defensive substances synthesized by plants are toxic to consumers is yet to be proved.
Moreover, source of transgenic has no relevance in assessing the toxicological aspects of food
derived from GM crops (Ramesh et al., 2005). One should be clear that it is easy to control
products of genetic engineering than that of traditional breed crops. Several closely related
molecules are obtained from the same substrate depending upon the metabolic state of the cell.
And, similar compounds often pose different biological activity. Therefore, potentially harmful
metabolites that may accumulate in GM plants as the result of unintended unpredictable effects
 Problems, Pitfalls and Advantages of Transgenic Crops 341

need to be carefully examined with an optimal balance between accuracy and coverage of
metabolite measure (Rischer & Oksman-Cladentey 2006). To avoid unintended unpredictable
effectsof GM crop products,a carefulintegrationof the data obtainedwith systembiologyis .
viewed as the only way that can lead to predictive engineering. Adequate regulations, constant
monitoring and further research are essential to avoid possible adverse effects of GM technology
to make it a safe option for consumers.

5. BRINGING WASTBLANDS UNDBK CULTIVATION

Salinization, alkalinization and desertification of lands are viewed as substantial threat for the
decline in total global area of cultivable land. Under such conditions, majority of plant species
are unable to establish themselves because of the limitation of water, nutrients, excess of salt
and high or low temperature. Among several approaches to improve degraded environments,
use of transgenic plants seems most promising.
Drought and salinity are correlated with each other and there are predictions that the problem
will become more severe in times to come. As if the current trend of climatic changes continues,
aquifers will be drawn down and competition between agricultural and non-agricultural water is
likely to be intensified (vide Pan et ai., 2005). For plants, to cope with salt and drought stress,
it is important to maintain vacuolar volume and the ion concentration within, by excluding Na+
at the plasma membrane or sequestering Na+ in the large intracellular vacuoles. The process is
controlled by H+ transport activity of vacuolar proton pumps. Out of three proton pumps operating
in plants, (P-type ATPase, vacuolar H+-ATPase and vacuolar H+ pyrophosphatase), two are
vacuolar pumps. Theoretically, enhanced expression of either of the vacuolar proton pumps will
increase the sequestration of ions in the vacuole by increasing the availability of protons and
vacuolar solute accumulation (Gaxiola et ai., 2001). Waditee et ai. (2002) reported that
overexpression of a Na+/H+antiporter genes in fresh water cyanobacterium enabled it to grow in
seawater. Furthermore, overexpression of AtNHX1, a Na+/H+antiporter gene in Arabidopsis plant
promoted its sustained growth and development in soil watered with 200 mM NaCl (Apse et
ai., 1999). The AtNHX1, a Na+/H+ antiporter gene is capable of catalyzing low affmity transport
of both Na+ and K+ (Vemema et ai., 2002). Similarly, membrane bound Na+/H+antiporter encoded
by 8081 gene plays important role in salt tolerance, as its overexpression improved salt tolerance
ability of Arabidopsis (Shi et ai., 2002). ( .
Control of ion flux activity by genetic manipulation of the ion transporters is I. nov~ and 01 j~
holds promises for the reclamation of farmlands lost to salinization and lack of rainfaii (GaxIOla et
ai., 2002). In addition, combination of the pump with various transporters may offer both diversity
and amplification of the stress resistant effects. Sergeeva et ai. (2006) reported that the expression
of bacterial 1-anlinocyclopropane-1-carboxylate (ACe) deaminase gene enhanced salt tolerance in
Brassica napus cv. Westar by decreasing the synthesis of stress ethylene. Besides, other strategies
adopted by plants to overcome water and salt stresses include; accumulation of osmoprotectants
(glycine, betanie and proline) and glyoxalase I (gly I) and glyoxalase II (gly II) enzymes involved
in the glyoxalase pathway (Singla-Pareek et ai., 2003). Freshwater cyanobacterium 8ynechococcus
sp. PCC 7942 transformed with E. coli Bet gene produced glycinebetaine and acquired resistance
to salt stress (Nomura et ai., 1995). Similarly, expression of N-methyltransferase gene from a
cyanobacterium Aphanothece haiophytica in Arabidiopsis, caused accumulation of high level of
 342 Advances in Life Sciences

betaine in root, stem, leaves and flowers and improved seed yield under stress condition (Waditee
et al., 2005). The dehydration response element (DRE), a cis-acting promoter element has
importance in regulating gene expression in response to environmental stresses. In transgenic plants'
overexpression of the transcription factor DREB lA interacts with the DRE and activates the
expression of many of these stress tolerance genes (Kasuga et al., 1999).

6. TRANSGBNICS AND GBNB f'WW

Gene flow leading to escape of traps genes is a potential concern associated with the introduction
0/j )b of transgenic cultivars. This includ change in the fitness of the wild relatives, concomitant risk
of increased weediness, loss of h~rbicide resistance to protect the crop ITom closely related
weeds, effects on genetic identity and diversity of sexually compatible relatives (Snow 2002).
Occurrence of gene flow among cultivars and their wild relatives is a multi-step process including
presence of cultivars or wild relatives within pollen or seed dispersal range, their ability to
produce viable and fertile hybrids, at least partial overlap in flowering time, actual gene flow by
pollen or seed, and establishment of crop genes in the domesticated or wild recipient population
(Gepts & Papa 2003). Moreover, chances of a trans gene released ITom a transgenic source
population to get established in sink populations depends on a number of parameters such as;
magnitude of the selective advantage, migration rate, genetic drifts, epistatic effects, and genotype
and environment interactions. These parameters are similar to those governing the fate of gene
flow in non-transgenes except, novelty of the transgenes in the case of former. Therefore, it
becomes very difficult to make predictions especially, when little is known about biological and
ecological effect of transgenes in their new genetic and environmental backgrounds. Ellstrad
(2001) believes that movement of unwanted crop genes in the environment may pose more of a
management dilemma than unwanted chemicals. For instance, single molecule of DDT
[1,1,ltrichloro- 2, 2,- bis (P-chlorophenyl) ethane] remains a single molecule or degrades, but a
crop allele has the opportunity to multiply itself repeatedly through reproduction thus, can Uustrate
attempts of containment.
Models predicting gene flow are largely assumptive and based on questionable premises
(Gressel 2005). Majority predicts for the occurrence of genetic swamping (Haygood et al.,
2003). However, following points invalidate the applicability of theses models.
1. In order to achieve the level of swamping, the wild relatives and the crop would have to
live in the same ecosystem (Haygood et al., 2003; Gressel 2005). However, there exists
a typical geographic separation between agro- and wild ecosystems. Flow of wild pollen
through wind current or insects decreases exponentially with distance usually to a low
asymptote. In a wild ecosystem the number of wild pollen is likely to out number the
crop pollens, thus reducing the chances of hybridization by crop pollen. AI-Ahmad et al.
(2005) in a replacement series experiment observed that with recurrent selection the
unfit genes gradually disappear. This further weakens the assumption of crop pollen
swamping wild type.
2. The assumption of synchronous flowering without self fertilization, but cross fertilization
in absence of any barrier is very much against the phenomenon of speciation. It is
exceedingly rare for crop pollen to fertilize another species but not the wild relative and
that too without any genetic barrier (Gressel 2005).,
 Problems, Pitfalls and Advantages of Transgenic Crops 343

3. Models assume for animal type replacement rates where a few progeny per mating is
typical, allowing lower fitness to become fixed. In contrast, most wild relatives of cr-ops
produce copious amount of seeds majority of which germinate in the area occupied by .
. the parents, ensuing the process of competition and eliminating less fit individuals.

Various strategies for the containment of gene flow have been developed (Table 1). For
example, Zhang et al. (2005) reported that a 50 m buffer zone can prevent the flow of genes
from transgenic cotton to surrounding area (Fig. I). Unlike drugs or pesticides plants have
never been subjected to a risk analysis. The very apprehension that antibiotic-resistance genes,
which sometimes remain in the transgenic crops, could spread to the pathogens and make them
less susceptible to the antibiotics has unanimously been ruled out by experts (Ellstrad 2001).

Table 1: Strategies for transgene containment.

Strategies Main tenet Efficacy Assessment

(a) Physical The main goal is to Keep away
transgenic pollen from physically
interacting with compatible plants.
(i) Avoidance Prohibits cultivation in areas where
chances of potential hybridization
wild species occur.
(ii) Establishment of Objective is to attain spatial separation.
suitable Buffer Thus reducing the chances of
zones transgenes spread.
(b) Genetic Contrary to conventional nuclear
method, it involves transgene
(i) Chloroplast incorporation into chloroplast. Since
engineering these are not transferred into pollen
grains, hence restricts the transgene
to the chloroplast genome, thus
preventing pollen mediated gene flow.
(ii) Mitigation- Involves engineering a mitigation gene
Gene approach along with the transgene. The mitigation
gene is show design so as to counteract
any fitness advantage of a transgene in
hybrids. It prevents the hybrid from
competing with wild plants and setting
seeds (AI- Ahmad et al. 2004).
Highly susceptible to human errors.
Probability of paternal chloroplast
transmission in tobacco in which transgenes
cannot be transmitted through pollen to
produce interspecific hybrid has been
observed to be 0.07% (Medysegy et al.
1986).
Mitigation gene approach has been
predicted to delay transgene escape for at
least 3000 generation in Rice having
recombination rate 1cm = 250 kb (Lee and
Natesan 2006).
Numerous 'domestication' traits in crop
plants that are capable of acting as
mitigation factor exist distributed throughout
the genome and are unlikely to be linked
genetically to a transgene. But possibility
of dissociation of these traits from the
transgene in hybrid persists because of
selective pressure in the natural
environment.

(iii) Genetic use
restriction
technologies (GURT)
Modificatipn of terminator technology;
based on repressible systems in which
seeds produced are non-viable unless
the plants are exposed to a specific
activator molecule stimulus
(Kuvshinov et al. 2004).
Most effective among all strategies available-
Needsto be used in combination with other
approaches for containment of transgene
conferring enhanced Chiotic stress
tolerance. Because they require different /2
management strategy because of much
greater selective advantages than other
modified traits.
344 Advances in Life Sciences

Fig. 1. A transgenics-planted field surrounded by elephant grass (about 4 m in height) to prevent gene flow
(International Center of Tropical Agriculture, Colombia; courtesy Prof. Tetsuko Takabe).

7. ENVIRONMENTAL RISKS ASSOCIATED WIT" EVOLUTION OF

RESISTANCE

The argument that deployment and widespread use of transgenic Bt resistant crops would lead
to the rapid evolution of Bt resistant insects needs empirical proof. Till date, resistance to Bt
crops in fields has not been reported, albeit laboratory selection has generated Bt resistant strain
in pests (Ferre & Van Rie 2002; Tabashnik et al., 2003). In contrast, external application of Bt
toxin as sprays resulted in resistance development in the field population of Diamond-back
moth Plutella xylostella (L) and green house populations of cabbage loopers Trichoplusia ni
(Hubner) (Janmaat & Meyers 2003).

7.1 Physiology and Genetics of Resistance

Morin et al. (2003) reported that in laboratory selected stains of pink bollworm insect fed on Bt
cotton survival depended upon three mutant alleles (r!, r2 and r3) of cadherin gene (BtR), which
confers resistance against CrylAC toxin. Each (r) allele is responsible for deletion of at least eight
amino acids upstream of the putative CrylAC- binding region of cadherin protein. However, Baxter
et al. (2005) reported that resistance to CrylAC in some str~ins of diamondback moth is not linked
with cadherin alleles. Though empirically unconfmned, Cerda & Paoletti (2004) believe that the
resistance is recessive and only one locus is involved. Till date, no resistant population has been
reported from the fields even in the case of monoculture, which experiences high selection pressure
(Tabashnik et al., 2006). Bates et al. (2005) observed that no resistance evolved in cotton and
 Problems, Pitfalls and Advantages of Transgenic Crops 345

maize, even when fanners did not comply with the modeled recommendations for refuges though,
resistant populations should be expected to appear in just a few years.

7.2 . Why are the Predictions Wrong?

Bt toxin affects more than one target sites on insects intestinal mucosa (Gressel 2005). Evidences
indicate that Bt CrylAC used in transgenic maize and cotton binds to both cadherin protein
(Flannagan et al., 2005) and aminopeptidase N (Bravo et al., 2004) with varying affinity. Multisite
binding has also been demonstrated for Cryl C toxin (Avisar et al., 2004). If two different
proteins bind, it is likely that either could mutate to non-binding. In that case, more than one
gene would be required to mutate, whereas, all the models assume single gene mutation at a
much higher frequency (10-3) than that observed wit;h mutation in non-insects (10-6) (Gressel 5) .
2005). An immunological study on the European cornborers with the cadherin receptors sug~est-=l ~~ f
.ul}
/~1 ~
.
inducing resistance a family of cadherins genes would have to. mutate. If so, then in case of
thigh level resistan'ce being recessive the resistance would be at an exceedingly low freq)l~ncy
(vide Gressel 2005). .

The possible explana!ion of resistance developing d~e to the application of Bt spray be

because of ipsects having single target receptor site. Kain et al. (2004) found that Bt resistance
in the cabbage looper was inherited as a single incompletely recessive gene. Strategies are being
developed by using binding site information to design the optimal mixture of genes which will
ensure that only few targets on each insect are affected, thus further lowering the possibility of
resistance evolution (Rang et al., 2004).

7.3 Fitness of Resistant Individuals

Resistant and susceptible organisms grown separately have identical productivity and fecundity.
However, in competition, resistant individuals disappear (Gressel 2005). This emphasizes for a
need to perform fitness experiments in wild; with resistant individuals in competition with the
wild type. Furthermore, the possibility of leaves having compounds that could reduce the fitness
of resistant individuals cannot be overruled. Unfortunately certain environmental conditions in
the field affecting fitness cannot be mimicked in the laboratory. Increased resistance to Bt sprays
in the field, greenhouse and laboratory selected strains of several major pests (Tabashanik et al.,
1990; Janmaat & Myers 2003; Tabashanik et al., 2003) still awaits definite answer.
However, the major concern regarding resistance to Bt crops is not of 'if' but 'when'. Even
though the researchers have begun thinking for second generation of insect resistant transgenic
plant. Avoidance of unnecessary exposure to the toxins still remains the most effective way to
delay resistance. However, this does not translate to a de facto moratorium on research to achieve
sustainability in insect resistance rather, should be taken as a call for exploration of more bold and
daring strategies. For example, use of inducible promoters could allow time and site specific
expression ofBt toxin (Cao et al., 2001) however, the effectiveness of plant-promoter system as a
resistance management tool remains to be established. Alternatively, novel Cry toxins with dissimilar
target sites (e.g., Vips) and non-Cry toxins with different modes of action not allowing cross
resistance to current Bt toxin (e.g., toxin A) are the other potential strategies besides Bt transgenics.
 346 Advances in Life Sciences

Vegetative insecticidal proteins (Vips) are produced by B. thuringiensis during vegetative growth
and are structurally and functionally dissimilar to their crystalline counterparts. They show
insecticidal activity against a wide range of lepidopteron and coleopteron pests (Lee et at., 2003). .
Similarly, toxin A, an insecticidal protein isolated from symbiotic bacterium Photorhabdus
tuminescens associated with entomopathogenic nematodes causes mortality during nematodes
infection. A key advantage of these toxins is that unlike Bt Cry toxin they have potential to control
unrelated pests (Bates et at. 2005 and references within). Use of fusion proteins is another strategy:
for example, combining the endotoxin CrylAc with the galactose-binding domain of the non-
toxic ricin B chain (RB). This fusion provides the toxin additional binding domains therefore,
increasing the potential number of interactions at the molecular level in target insect. Such fusion
proteins (e.g. Cryl Ba/Cryl Ia and BtRB) provide strong and sustainable resistance as they require
multiple counter adaptive mutations (Naimov et at., 2003; Mehlo et at., 2005).
Nevertheless, despite the continued success of Bt crop~many believe that resistance
cI>! remains a threat. To mitigate this, an integrated resistance management practice involving the
most effective strategies such as high-dose-refuse and stacked genes together with biological
and cultural control will have an edge over adoption of a single strategy. This will also reduce
the inevitable evolutionary eventuality of recurrence of resistance. Monitoring resistance
(Table 2) in field populations has an important role in formulation of future integrated resistance
management strategies. Efforts are needed to evolve specific resistance management strategies
on case to case basis.

Table 2. Techniques involved in monitoringof resistance.

Techniques
Diagnostic or discriminatingdose incorporation
method (cf. Bates et al., 2005)
Main tenet
Involves incorporationof diagnostic or discriminatingdose
into an artificialdiet. Such dose allows only resistant
individualsto survive. Technique is relativelyinexpensive
enabling detection of both polygenic and multiple
resistance mechanisms Major concern is its inabilityto
detect alleles, which are recessive or occur at low
frequency.

F2screen method (Andow and Alstad 1998) Capable of detecting rare recessive alleles; but fails to
detect polygenic resistance.

DNAbased screening method (Tabashnik et al., 2006) Highlyefficient than conventional methods used earlier.
Involves use of PCR based primers, and is capable of
detecting even single resistance alleles in heterozygotes.

8. REDUCING PESTICIDES USAGE AND POLLUTION

Pesticides are the major threat to the environment and human health. Every year, farmers use
2.54 billion kg of pesticides amounting $33.5 billion for controlling pests (vide Pan et at.,
2005). Major problems associated with the continued and excessive use of these chemicals are
the persistence of their residues in foodstuffs, pollution of underground water resources, shift in
microbial diversity, development of resistance among pests and harm to non-target organisms.
The toxic residues of pesticides and herbicides in foodstuffs cause health problems such as
deficiency of immune system and cancer in exposed population. Planting transgenic pest-resistant
 Problems, Pitfalls and Advantages of Transgenic Crops 347

plants can reduce or even eliminate the use of these chemicals thus, minimizes environmental
damages (vide Pan et al., 2005).

8.1 . Monitoring, Detecting and Detoxifying Environmental Pollution

Contamination of natural resources is a main concern of the modem world. So is their monitoring,
detection and detoxification. In recent time, synergy between molecular biology and genetic
engineering has added new dimensions to this area. In environmental monitoring, a shift from the
classical organism level markers to biomarkers with a high degree of sensitivity, efficiency and
reproducibilityhas taken place (Monciardiniet al., 1998). Biomarkers are proteins encoded by
specific genes under specific environmental conditions. Creation of transgenic plants with specific
genetic biomarker has widespread applications as they release signals encountering specific
environmental change. For example, transgenic tobacco plant transformed with the plasmid
constructed by fusing stress inducible promoter of the barley gene Hvhsp and reporter gene Gus,
showed enhanced Gus expression when exposed to heavy metals (Monciardini et al., 1998).
Besides being expensive conventional techniques of remediation often prove insufficient (Dixon
1996). Phytoremediation involves use of selected or engineered plants for environmental clean up,
and is applied in situ without affecting the texture and structure of polluted soil (Heitzer & Sayler
1993) (Table 3). For heavy metal removal, phytoremediation has following subsets:
1. Phytoextraction: This involves the use of metal accumulating plants that can transport and
concentrate metals from the soil into the harvestable parts of roots and above ground shoots.
2. Phytovolatilization: In this process, plants extract volatile metal (e.g., mercury, arsemic,
selenium) from soil and volatilize them through foliage.
3. Rhizofiltration: In this case, plant roots absorb, precipitate and concentrate toxic metals.
. 4. Phytostabilization: The process involves the use of plants to eliminate the bioavailability
of toxic metals in the soil.

Table 3. Strategies involved in phytoremediation of metals.

Approach Gene(s) Plant Pollutant Reference
manipulated
Overexpression of MTI and MT2 genes Tobacco Cd Pan et al., 1994
genes encoding encoding for Oil Seed rape
metal chelating metallothionein
'--- proteins or metal
t binding enzymes ICUP1 gene from Tobacco Cu Thomas et al., 2003
yeast
Overexpression of ZNT gene encoding Arabidopsis Pb/Cd/Zn Lee et al., 2003
metal transporters for Pb (1I)/Cd(II)1
genes Zn(lI) pump
CAX2 Tobacco Ca2+/Cd2+/Mn2+ Hirschi et al., 2000
Grafting metal Mer A gene Arabidopsis thaliana Hg2+ Rugh et al., 1996
metabolism path a,s C Cd Dhankher et al., 2003
way from other
organism into plant

Producing antibody Deaminase gene Tomato/Canola Cd, Co, Cu, Mg, Grichko et al., 2000;
or some stress Ni, Pb, Zn Nie et aI., 2002
induced enzymes
using transgenic

-,
348 Advances in Life Sciences

During the. process of metal uptake, translocation and sequestration several genes are
involved. Overexpression of any or combination of these genes is the best possible strategy
for creating transgenic plants for phytoremediation of toxic metal (Pilon Smith & Pilon'
2002). Moreover, besides metal and organic pollutants, transgenic plants can also be used
to degrade explosives (e.g., TNT, RDX, nitroglycerine) or other xenobiotic compounds. For
example, plants transformed with genes of soil bacteria Enterobacter cloacae PETN reductase
were capable of degrading explosives (Rosser et al., 2001). Tobacco plants containing
mammalian cytochrome P450Zel were able to degrade wide range of pollutants such as
TCE, ethyl bromide, carbon tetrachloride, chloroform and vinylchloride (Doty et al., 2000).
Shimizu et al. (2002) reported the expression of catechol degrading gene cbn-A in plants
that encodes the enzyme involved in the cleavage of the aromatic ring of chlorocatechol.
Second generation approach of phytoremediation includes cellular targeting especially the
vacuoles and creation of artificial metal sinks in the shoot by enhancing metal binding sites
(Eapen & D'souza 2005).
At small-scale field experiments, there are number of native species used successfully for
the phytoremediation (Blaylock 2000). However, commercial use of trans genies for
phytoremediation has yet not been possible (Eapen & D'souza 2005). Large scale field testing
of transgenics under normal environment has yielded encouraging results. For examples; nearly
200 popular trees containing modified MerA gene planted on a mercury contaminated site of an
abandoned hat factory in Danbury CT, successfully volatilize Hg(O) (Rugh et al., 2000). Similarly,
overexpression of Cystathione-y-synthase in Brassica juncea enhances volatilization of selenium
(Se) (Huysen et al. 2003). CommerGial application of phytoremediation technology is feasible
only when accumulation approaches 100 to 1000 folds to that of the wild type. Unfortunately,
this still remains unattainable. Perhaps the most critical challenge for the field is to develop
plants having superior ability of contaminant accumulation witp. near normal biomass. In addition,
overexpression of the same gene in different species has exhibited different effects like increasing
tolerance in one and sensitivity in the other (Thomas et al., 2003). Other limitations that hamper
the efficient application of phytoremediation include; time required for acceptable effects, limited
depth of the root system, slow growth of plants, sensitivity towards some pollutants, problem of
being part of a food chain and dependence on changes in the climate and winter dormancy
(Khan et al., 2000).
A possible risk involved with transgenics mediated phytoremediation is enhanced exposure
of wild life and humans. However, the risk of metal ingestion by wildlife during browsing/
grazing can be prevented by suitable fencing off the area, and use of non-palatable species.
Studies indicate that mercury and selenium volatilizing plants pose negligible threat to the
environment (Rugh et al., 2000; Lin et al., 2000). There is an urgent need to design and
develop trans genies for specific uses such as targeted remediation of a particular metal e.g.,
arsenic in Bangladesh. This can only be achieved if the strategies are multipronged, target
specific, and financially viable for its wide spread applicability and acceptability. Integration
of modeling techniques. with data on genome, transcriptome, proteome and metabolome is
most likely to enhance our ability to predict the effects of transgenes on contaminants uptake
and tolerance.
 Problems, Pitfalls and Advantages of Transgenic Crops 349

CONCLUDING REMARKS

Debate on genetically modified plants is neither new nor surprising. It arises from conflicting'
values and priorities, the perceptions of past technical abuses and rational concerns. However,
rather than based on sound scientific knowledge. Most of the criticisms are largely philosophical
and conjecturaL Extended field based studies could provide a clear message about perceived
advantages and disadvantages of transgenics. Till date, there has been little evidence to support
either case. Further, research is needed to determine the true safety of these plants and products
for environment and population that consume these products.

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