Transfer of herbicide resistance genes via pollen-mediated gene flow
from genetically engineered (GE) crops to non-GE crops is of relevance
The transfer of herbicide resistance genes via pollen-mediated gene
flow from genetically engineered (GE) crops to non-GE crops is of relevance
for ensuring co-existence of different agricultural cultivation forms as
well as for weed management. Co-existence in oilseed rape (Brassica napus)
depends on the development of management strategies to keep the adventitious
presence of GE plant material below the EU labeling threshold of 0.9% in
non-GE harvest products. Crop-to-crop cross-fertilization is one source of
adventitious GE presence. Several field experiments have been conducted to
evaluate pollen-mediated intraspecific gene flow from herbicide resistant to
nonresistant oilseed rape. We have performed a literature search for
worldwide studies on cross-fertilization in oilseed rape1 to identify the
major factors affecting pollen-mediated gene flow.

Pollen-mediated gene flow in oilseed rape
Most of the studies investigated (n = 16) could be categorized as
having either a continuous design (n = 7), in which a donor plot is
completely surrounded by receptor plants, or a discontinuous design (n = 9),
in which the receptor field is on only one side of the donor, either
adjacent or at a distance. Studies using individual fertile plants as local
pollen traps to measure gene flow were not considered because due to the
absence of local pollen competition they do not provide any information on
outcrossing rates under the conditions of agricultural production.

Figure 1 shows the mean values of cross-fertilization for continuous
and discontinuous design trials at several distances based on all studies in
which average outcrossing data were available. Using the continuous design,
the average values of cross-fertilization are highest immediately adjacent
to the source (1.78% ? 2.48) but are frequently constant around 0.05% (?
0.05) over tens of meters. For discontinuous field trials, the outcrossing
rate declines slowly and steadily from a mean value of 0.94% (? 0.51) next
to the source and is constant around 0.1% (? 0.11) over a hundred meters. In
general, all studies demonstrate a steep decline in cross-pollination rates
with increasing distance and that the bulk of cross-fertilization occurs
within the first 10 m of the field. However, various biological and physical
parameters, e.g., size, shape, and orientation of the pollen source and the
recipient field, isolation distance, wind characteristics, rain, local
environment, genotype, and zygosity, influence cross-fertilization in
oilseed rape.




Shape, orientation, and size of pollen source and recipient field
A continuous design seems to favor short distance pollen dispersal; a
clear and high edge effect and a rapid decline of cross-fertilization over
the next 50 meters is observed. Most field experiments in this design class
used small transgenic plots and relatively wide nontransgenic border areas.
It has been observed that large sink populations subsidize the species pool
of small source populations via a mass effect. In general, the larger the
recipient field compared to the donor field, the lower the probability of
cross-fertilization. In the discontinuous design field trials, pollen
pressure from both donor and recipient is presumably equal because of
similar field sizes. As a consequence, the outcrossing rate in this design
class declines slowly and steadily over the first tens of meters and levels
off at around 0.1% over a hundred meters. Not only the size but also the
alignment of donor and recipient fields is important for levels of
outcrossing. With a donor field size of 2 ha, the rate of outcrossing might
be much higher than from a 10 ha field, if the long side of a recipient
field is facing the source field as compared to the short side. In other
words: the deeper the recipient field, the lower the cross-fertilization
level of the total harvest product.

Isolation distance and border crops between pollen source and
recipient field
Isolation distance is one means to ensure seed purity (e.g., 100 m for
certified rapeseed). Some outcrossing studies investigated the effectiveness
of isolation zones for reducing gene flow compared to the use of
nontransgenic buffer areas. When crops are isolated by open ground or low
growing crops, it appears that the first rows of the recipient field
intercept a high proportion of foreign pollen due to the low convarietal
pollen load of the field margin. When there is no gap between transgenic and
nontransgenic fields, the plants located in the contact area act on one side
as a pollen trap; on the other side, plants produce additional pollen that
dilutes the transgenic airborne pollen, so that at within-field distances
comparable to a certain isolation distance lower rates of
cross-fertilization are observed.

Genotype and zygosity
Different herbicide resistant plant varieties, in particular with
resistance to glufosinate and glyphosate, have been used in outcrossing
studies. Each transgenic variety can show different levels of outcrossing
under the same experimental and environmental conditions, influenced by
differences in flowering time, pollen quantity, and selfing rate. In
addition, in the case of homozygous glyphosate and glufosinate resistant
plant lines, all pollen carries the herbicide resistance gene. By contrast,
in studies of cross-fertilization from glufosinate resistant hybrids, the
amount of transgenic pollen was lower, resulting in only about 5/8 of the
outcrossing frequencies of homozygous herbicide resistant lines. Moreover
when measuring outcrossing via herbicide spray tests, one has to consider
that gene dosage effects have been demonstrated in several cases by
comparing hemizygous and homozygous transgenic plants with homozygotes
usually having higher transgene expression levels. Therefore hemizygous
seedlings with low expression levels of the herbicide resistance gene might
not always be easily distinguishable from unmodified susceptible plants.

Local environment and climatic conditions
The range of cross-fertilization at a given location is also
determined by the narrow range of weather conditions and local topography
around the field trial site, and the numbers of bees and other insects that
are likely to increase the amount of pollen transfer.

Management strategies to reduce gene flow
Due to additional sources of adventitious GE presence?seed impurity,
volunteers, adventitious seed transfer during harvest and transport?a
maximal value of 0.5% for crop-to-crop cross-fertilization is relevant
within the 0.9% threshold for the adventitious presence of GE crop products
in nontransgenic food and feed set by EU labeling legislation. Technical
measures for achieving co-existence have to ensure that thresholds will not
be exceeded on a long-term basis. The first few oilseed rape rows intercept
a high proportion of pollen when open ground or low growing barrier crops
separate oilseed rape fields. The removal of the first 10 m of crop along
the side of a nontransgenic field facing a GE crop might be more efficient
for reducing the total level of cross-fertilization in a recipient sink
population than to recommend separation distances. It appears that the use
of predominantly self-pollinating, male sterile, or cleistogamous cultivars
as a biological containment strategy will also reduce gene flow2,3.

The adventitious presence of GE oilseed rape is not only affected by
outcrossing via pollen-mediated gene flow, it is also affected by volunteer
populations within fields via seed-mediated gene flow. The dynamics and
persistence of volunteer oilseed rape is mainly influenced by field
management practices (time of first tillage following oilseed rape, cultivar
type, tillage depth, crop rotation)4. If volunteers flower,
cross-pollination to other oilseed rape plants and fields can occur.
Herbicide resistant volunteers arising from unintended gene flow can be
easily managed with herbicides if the subsequent crop is a
non-herbicide-resistant cereal5 but will require special weed control
strategies in the case of crops with resistance to the same herbicide. In
order to avoid the formation of multiple herbicide resistant plants, farmers
should not grow cultivars with different herbicide resistances in adjacent
fields.

Feral populations are widespread at relatively low densities in
regions cultivating oilseed rape. Pollen flow from sporadic occurrences of
feral oilseed rape to neighboring rapeseed fields can be considered a rare
event due to the high amount of competing field pollen. Therefore feral
plants as a source for further transgene flow may only be a realistic
scenario if large feral populations are present near an oilseed rape field.
Nevertheless, volunteer and feral population dynamics should also be taken
into account when assessing sources for adventitious GE presence and
feasibility of coexistence.

References

1.. Hüsken A, Dietz-Pfeilstetter A (2007): Pollen-mediated gene flow
from herbicide-resistant oilseed rape (Brassica napus L.). Transgenic Res.
16, 557-569
2.. Devos Y, Reheul D, Schrijver A, Cors F, Moens W (2004)
Management of herbicide-tolerant oilseed rape in Europe: a case study on
minimizing vertical gene flow. Environmental Biosafety Research 3, 135-148
3.. Pierre J, Fargue A, Picault H, Pinochet X, Renard M (2007).
Methods to study advantages of cleistogamy in oilseed rape in limiting
unwanted gene flow. Proc. of the 12th International Rapeseed Congress,
Wuhan, China, March 26-30, 2007. vol.1:177-179
4.. Gruber S, Pekrun C, Claupein W (2004) Population dynamics of
volunteer oilseed rape (Brassica napus L.) affected by tillage Eur. J.
Agron. 20, 351-361
5.. Downey RK (1999) Gene flow and rape ? the Canadian experience.
In: Lutman PJW (ed.) Gene Flow and Agriculture ? Relevance for Transgenic
Crops, British Crop Protection Council, Vol. 72, pp.109-116


[www.isb.vt.edu]