Comment on

'Report on the separation distances required to ensure cross-pollination is below specified limits in non-seed crops of sugar beet, maize and oilseed rape'

published by the

Ministry of Agriculture Fisheries and Food,

Compiled by J.Ingram July 2000.

National Institute of Agricultural Botany, Huntington Road, Cambridge

by

National Pollen Research Unit,

University College Worcester, Worcester WR2 6AJ

September 12^th 2000

Introduction

Comments by National Pollen Research Unit ( NPRU) on the MAAF document.

NPRU has been asked by Friends of the Earth to comment on the MAAF document (Ingram 2000). In this report the points highlighted in the review are discussed in the light of the original references cited in the document as well as additional references, where relevant. It has not always been possible to give detailed comment on all aspects of the review as some references are not yet in the public domain or full details of references are not given.

Format of NPRU comments

The review by Ingram (2000) is considered on a point-by-point basis in the order of the original text. To set a context for the comments very brief summaries are made from Ingram and direct quotes are used where it is felt to be necessary. We would, however, recommend the use of the whole document for the best understanding of the comments made about it.

The Ingram report considers separation distances specifically to prevent cross-pollination but the subject, as with most biological themes, does not stand in isolation. Therefore, where particularly relevant , we have included brief details of other factors from the wider debate of gene flow from and to crops. Useful further references of such issues, including gene flow through seed, introgression into wild relatives and feral populations, can be found in Lutman (1999).

1. Objectives

Ingram (2000)

In the section on Objectives Ingram comments that, " After considering all the results available, a robust , representative set of data has been identified.".

NPRU comment

This is highly debatable as very few detailed studies have been conducted on cross pollination over a range of " typical farm situations ".

Section 2 - Sources of information

Ingram (2000)

The Ingram report outlines the potential pros and cons of different sources of information. The importance of the scale of experiments is acknowledged, in that results derived from normal scale agricultural plots are likely to be a better model than results from smaller, experimental plots. Also highlighted (2.4) is the potential effect of varying environmental conditions/experimental error on the results between experiments.

Section 3 - Factors affecting the degree of cross pollination

Ingram (2000):

The importance of a consideration of source scale in making any recommendations is noted.

NPRU Comment:

We would specifically add to this that different fields/ feral populations can combine on a regional level to elevate background levels of pollen (Squire et al. 1999). The number and distribution of plots is also relevant besides source scale. The importance of this was also noted by DETR (1999).

Ingram (2000):

The attractiveness of crops to pollinating insects is mentioned.

NPRU Comment:

The numbers, types and pollen preferences of insects, as well as the breeding system of the crop in question, are also vital factors in the role of insects in cross-pollination.

Ingram (2000):

The point is made that the importance of pollination vectors ( wind v. insect ) may be affected by environmental conditions and could differ between countries perhaps affecting the relevance of non-UK studies, although there is also variation more locally. The role of barriers in reducing pollen dispersal is noted.

NPRU Comment:

The doubts about the relevance of non UK studies illustrates the point that the available data cannot be considered to be a " robust and representative set" .

3.3 Factors affecting the degree of cross-pollination once pollen has arrived at the receptor crop

Ingram (2000):

Perfect synchrony between the flowering times of two crops would give the highest rates of cross-pollination.

Pollen viability is unlikely to be a limiting factor to cross pollination under normal conditions

The rate of cross-pollination from incoming pollen is dependent on the level of competition from pollen produced by the recipient crop itself. The example is given of modern varietal association with high levels of male sterility that are more prone to cross pollination with pollen from another field than are conventional fully fertile varieties.

NPRU General comment on section 3.0

This is a fairly balanced summary of factors affecting pollen dispersal albeit brief. Other factors could, however, have been noted including size of receptor plot (although it is considered in subsequent chapters): A small recipient block will have a higher proportion of cross-pollination than a larger one, all other factors being equal.

Some consideration should be given to the difficulties of predicting the responses of any agro-ecological system with time. For instance the species and/or numbers of pollinator may change. For example, 'unusual' behaviour in pollen beetles is likely to have led to high levels of cross pollination (31%) at 1km in a study on potatoes by Skogsmyr (1994). The importance of numbers and distribution of source blocks should also be discussed in the context of the influence on background concentrations of pollen.

Section 4. Maize and Sweet corn

4.1 Introduction

4.2 Practical experience of separation distance in maize.

Ingram (2000) report:

Descriptions of recommended separation distances from practical experience are given.

NPRU Comment:

The value of citing references for recommendations is limited without some direct reference to testing.

4.3.1 Results of Maize experiments

Ingram (2000):

Details of studies on maize pollen dispersal are described and in particular those of Jones and Brooks (1950).

4.3.2 Measurements of individual factors which influence cross-pollination

4.3.2.1 Pollen source

Ingram (2000):

The report notes the probable importance of emitter size, although no specific data is available for maize. Comment is made about the balance between horizontal and vertical movement of pollen and the fact that vertical distance between adjacent crops might affect the amount of pollen that passes to the receptor crop.

NPRU Comment:

Pollen from various taxa differs in dispersal characteristics, but in all cases an increase in the pollen cloud due to an increased size of source area is very likely to lead to a direct increase in cross-pollination as noted in 3.1 of Ingram (2000).

The vertical dispersion of pollen can be influenced by many factors including general turbulence in the air flow, local eddies due to barriers and topography and by convection currents. In some weather situations upward movement on air currents can be predominant, followed by lateral transport aloft and subsequent deposition at considerable distance downwind. Vertical differences in crop location could be important but it is also pertinent to mention that transport of pollen between crops at the same altitude may not be in the horizontal direction.

4.3.2.2 Transmission to recipient crop

Ingram (2000):

The report notes that cross pollination by insects is unlikely and discusses the role of barriers.

NPRU Comment:

Although cross-pollination via insects is improbable, the pollen collected by bees is likely to become a constituent of honey.

4.3.2.3 Factors affecting degree cross-pollination once pollen has arrived at recipient.

Ingram (2000):

The roles of competing pollen and its synchronisation are noted.

NPRU Comment:

Importance of area of the recipient field as well as orientation should be introduced here.

4.4 Factors affecting the degree of gene expression in the harvested crop after pollination by foreign pollen

A short section with factual content only

4.5 Recommendation on separation distances for maize and sweet corn.

Ingram (2000):

The recommendations are based on a number of assumptions including that the field is rectangular and at least 2ha. No allowance made for contributions due to contaminated seed or volunteer plants. Also the point is made that cross-pollination occur mainly in the rows of the recipient crop closest to the source.

Ingram (2000) state that 'The best body of data for estimating levels of cross pollination is that of Jones and Brooks but it represents a worst case scenario because of the high wind and the dry conditions prevailing during the experiment. Recent unpublished data also suggests that [Jones and Brooks, 1950] showed more extreme pollination than is normal in France.'

NPRU Comment:

We concur that the Jones and Brooks (1950) study is the best body of data currently available we do not agree that it represents a worst case scenario. The report itself (Section 4.3.1, Ingram, 2000) states that 'There was considerable difference in the crossing between years, one wet season with less wind than usual, roughly halving the crossing rate.' The varied weather conditions over the three growing seasons clearly do not correspond to 'high wind and the dry conditions prevailing during the experiment'. Furthermore, in the original Jones and Brooks (1950) paper there is no mention of high wind conditions.

To use an unpublished study to re-enforce a point without giving any source details is irregular in scientific correspondence,(Quote from Ingram "recent unpublished data also suggests that the American study showed more extreme pollination than is normal in France"). We cannot make any comment on the interpretation of this particular unpublished study as no details are given.

Ingram (2000):

A table and text is presented giving recommended separation distances "based on data of Jones and Brooks".

NPRU Comment:
No indication is given of what part(s) of the data of Jones and Brooks have been used/omitted in the calculations. No method is supplied for the calculations either as part of this report or as a reference to other work. It is highly unusual for recommendations to be proposed on such a vague basis.

Detailed comment on the adequacy of the proposed separation distances is not possible. However, Jones and Brooks (1950) themselves question the adequacy of the then newly imposed isolation distance of 200m. , '…..40 rods isolation may be insufficient to produce seed of the degree of purity required by the international standards, i.e. a maximum of 1 percent of other varieties'. In the summary/conclusion they state 'The percentage of outcrossed seed was greater than the maximum mixture permitted by the international standards at distances of 40 rods isolation, or less. In one of the three years the outcrossing exceeded this maximum at 60 rods [=300m]'

Table 1a shows the summarised results from Jones and Brooks (1950). The mean outcrossing rates for the three years of the study is 1.2%, while for one year (1949) it is 2.5%. It is significant to note that this figure represents the mean for the whole of the square recipient block. Had the field been rectangular with its long edge facing the source the degree of cross pollination is likely to be higher due to the higher relative levels of cross-pollination in rows closest to the source. To illustrate the higher level of cross pollination in rows adjacent to the source field in 1949, at 200m separation distance, the mean of the first five rows was 7.1% (range 14 - 0.7 % cross-pollination).

No allowances are made in the recommendations for the size of pollen source. The importance of this has been described in section 3.1 of Ingram (2000) 'There does seem to be an a priori case for increasing emitter size produce a larger pollen cloud and thus all other factors being equal to lead to increased pollination.' In other words a recipient field (of a given size) would have increased levels of cross-pollination from a nearby large field than a small one.

Table 1 a,b: Maize pollen dispersal. a) Average percentage of outcrossing

Jones and Brooks (1950); b) Percentage outcrossing in between two fields of maize recorded for 10 distances After Salmov (1940) cf. Jones and Brooks (1950).

Distance north of source fields (m)

Year	0	25	75	125	200	300	400	500
1947	35.1	16.5	5.1	0.8	0.4	0.2	0.2	0.2
1948	17.9	7	3.6	2.5	0.7	0.3	0.2	0.1
1949	32.9	19.2	8.6	3.7	2.5	1	0.3	0.3
Mean	28.6	14.2	5.8	2.3	1.2	0.5	0.2	0.2

b. Distance from pollen source (m)	10	50	100	150	200	400	500	600	700	800
Mean hybridisation percentage	3.3	0.3	0.4	0.3	0.5	0.02	0.1	0.8	0.2	0.2

Heading 'Multiple adjacent fields'

Ingram (2000)

The report suggests that adjacent fields are unlikely to affect the level of cross-pollination above the threshold level.

NPRU Comment:

No references are given to support the statements given.

With large scale planting it is likely that the background level of pollen will be increased on a regional level, in effect increasing the source scale (Squire et al., 1999). Even on a local level it would seem that fields in the direction opposite to that of the prevailing wind can still be a significant pollen source. Salamov (1940; cf Jones and Brooks, 1950), using a similar methodology to Jones and Brooks, showed a cross-pollination rate of 0.2% at a separation distance of 800m (Table 1 (b)).

General comments on section 4.

One of the main problems with this section is the lack of explanation of calculations and methodology for recommendations. The reliance on Jones and Brooks by Ingram (2000) and other reviews (e.g. Treu and Emberlin, 1999) demonstrates the very limited relevant data available for maize. This illustrates the need for further experimental research in a number of areas (including source and recipient scale and landscape effects) before any recommendations are adopted.

5.0 Oil seed rape

5.1 Brief description of oilseed rape and crop use.

NPRU Comment:

It may be worth noting that as well as seed being utilised, seedlings are often used in salad cress.

5.2 Practical experience with separation distances

Ingram 2000

The report gives the separation requirements for certified seed production (200-400m) and notes that only very rarely are the minimum purity levels (99.7-99.9% for conventional varieties and 90% for hybrids) not met.

5.2.2 Separation distances for HEAR crops

Ingram (2000):

The report notes that low-high erucic acid oilseed rape crops are not a suitable model for GM cross-pollination. [A 50m minimum separation distance of 50m exists between these two crops (DETR, 1999)]

5.3.1 Field experiments measuring the effect of separation distance on cross-pollination.

Ingram ( 2000)

Studies by Downey (1999) and Champolivier et al. (1999) are discussed.

It is noted that studies using a very small source scale are of limited value.

A study by Simpson (2000) highlights the fact that cross-pollination within varietal associations can be considerably higher than conventional varieties.

NPRU Comment:

The figures referred to in the report for Champolivier are not immediately apparent in the original conference paper. Only a graph is given: at c.2m a maximum of 6.9% cross-pollination was achieved (recorded as double resistance rate) while at c.112m a maximal cross pollination rate of c.0.5% was recorded.

We cannot comment on the data by Simpson (2000) as it is not yet published (this document should be cited as Simpson (unpublished)).

Conspicuous by its absence is the work by the Scottish Crop Research Institute, including Timmons et al. (1995, 1996) and Thompson et al. (1999). When considering gene flow between neighbouring oilseed rape crops DETR (1999) noted that the work of Timmons et al. (1995, 1996, in press) was 'Arguably the most relevant research…'

The most recent research by Thompson et al. ( 1999) shows that c.5% cross-pollination occurred at 4000m from the nearest field with bees as the likely vector. At the closer range of 100m an average of 61% was recorded. The results also showed the pollinations to be from a mixed source giving further evidence of the influence of elevated background concentrations due to multiple sources ( Squire et al. 1999). The research uses male sterile trap plots which are likely to overestimate the degree of cross-pollination, especially with fully fertile varieties. However, as the authors point out varietal associations or any other crop types with degrees of male sterility are more likely to be pollinated by external sources. This has been substantiated in the so called Advanta incident in the UK, where 'around 1%' of non-GM seed was found to be contaminated with a GM herbicide resistance trait. This is likely to have been caused by cross-pollination in the Canadian seed producing area. According to Advanta the source field was a minimum of 4000m from the recipient (House of Commons Agriculture Select Committee, 18^th July 2000). (Full report of the incident is awaited).

5.3.2 Measurements of individual factors influencing cross-pollination

5.3.2.1 Emitter crop - Pollen source

Ingram (2000):

Descriptions are given of the size of the pollen source and oilseed rape attractiveness to insects.

NPRU Comment:

The work of Osbourne et al. (1999) in fact concentrates on Bumble bees (Bombus terestris) rather than honey bees (Apis mellifera), as would appear to be implied by the report. In the study Osbourne et al. show that bumble bees forage far wider than had originally been thought and note that this may have implications for GM crop pollen dispersal.

Research by Ramsay et al. (1999) shows that worker bees in fact switch crops between foraging trips and/or mix pollen within the hive. The study concludes 'With most honey bee colonies foraging up to 2km from the hive, some pollen transfer up to 4km must be expected'. Also, noteworthy is that honeybee hives are often deliberately introduced by bee keepers.

Furthermore, consideration of other pollinating insects should also be made, especially the pollen beetle (Meligethes aneus). No studies would appear to be available of the role of these beetles in between crop pollination, although they are very common (see below).

5.3.2.2 Transmission to recipient crop

Ingram (2000)

The report notes that both wind and insects can play a role in oilseed pollination and discusses the possible effect of barriers. It is felt that long distance cross-pollination by bees between fields is not likely to occur at anything but trace levels.

NPRU Comment:

Please refer to the comments for 5.3.1 and 5.3.2.1.

5.5 Other evidence in oilseed rape cross-pollination

Ingram (2000):

Considers evidence from Simpson (1999 and 2000/unpublished) highlighting small levels of cross-pollination.

NPRU Comment:

As previously noted Simpson (2000) is not available as it is not yet published. Simpson et al. (1999) conclude ' Results from these initial studies of gene flow in oilseed rape show that pollen can be dispersed over considerable distances indicating that gene flow can occur from releases of GM herbicide tolerant oilseed rape'. Male sterile trap plots showed that in the direction of the prevailing wind (N and NW) show a pollination frequency of c. 27% at 100m, c.19% at 200m and c.7% at 400m (data read from Figure 2). These relatively high figures are despite a pollen barrier up to 20m wide and a smaller pollen emission due to some male sterile plants within the source. These factors must be balanced, however, against the very small plot size (6 plants) and their male sterility.

5.6.1 Recommendations on separation distances for conventional varieties of oilseed rape

Ingram (2000):

Recommended separation distances are given in tabulated and text form.

NPRU Comments:

It is not clear which part of the data set (Simpson 1999 and/or 2000) is being used or the method of calculation that is employed.

We can see no reason why the recommendations for separation distance certified seed crops should be any different for the normal oilseed crop as the potential for contamination through cross pollination is the same. For instance, for the long side of a 2ha field at a separation distance of 11.5m this is given in the report as 0.24%. Why, then, is the separation distance for certified seed with 99.7% purity at 200m for the UK? No allowances are made for the source scale either at local (i.e. size of source field) or regional levels (landscape scale effects).

In a DETR (1999) report on the environmental risks of herbicide tolerant oilseed rape (section 2.2.5) the authors conclude that:

' complete genetic isolation of oilseed rape, were it required, would have to be on a regional (tens of kilometres) scale, and that, under current farm practices, local contamination between crops (probably at a low, but also variable, level) is inevitable.'

5.6.2 Recommendations on separation distances for hybrids

Ingram (2000):

For varietal associations the report estimates that '1% cross-pollination between fields is probably achieved by 100m separation distance'

NPRU Comments:

This contrasts sharply with the studies cited above, as well as the recommendation for hybrid certified seed where a 10% cross-pollination (=90% purity) threshold is allowed for at a minimum separation distance of 300m.

NPRU General comments on section 5

The potential for gene flow in oilseed rape is a far more complicated issue than in a crop such as maize. In oilseed rape gene flow can also occur through other routes (for more details and references see Treu and Emberlin, 1999; section 3.0). There are other factors, although not as direct, that can affect cross pollination To omit these factors from any discussion on the potential for cross-pollination between crops is misleading giving an oversimplified picture.

i) Volunteers. Seed is very small (1000 seed weight is 5g; Lutman, 1993) and is therefore easily spilled in large numbers. This seed can be distributed in space via transportation potentially giving rise to feral populations (see below) as well as in time through establishment in the seed bank. Seed can remain viable in the seed bank for many years

ii) Ferals . Feral populations resulting from such are common in the UK (see for instance Crawley1993, Squire et al., 1999). As Feral populations are normally far smaller than are crop fields of oilseed rape they are particularly vunerable to cross-pollination from a field scale source, even at long distances (DETR, 1999). In turn feral oilseed rape has the potential to become a pollen source.

iii) Wild relatives. Data suggests that spontaneous hybridisation, can occur between oilseed rape and at least six wild species found in the UK (DETR, 1999; Treu and Emberlin, 1999). Many authors regard the introgression of transgenes into wild relatives inevitable with large scale GM releases, even with low rates of successful hybridisation (e.g. Ellstrand, 1988; Kereiva et al., 1994 and Leckie et al., 1993). Wild relatives may then also contribute to the GM pollen source, although the source scale of wild populations compared to crop fields is normally small. However, the habit of the very common (where uncontrolled) pollen beetle (Meligethes aeneus) may promote the frequency of wild-crop and crop-wild gene flow. In a study by Free and Williams (1978) pollen beetles were found abundantly on nearby early flowering cruciferous and non-cruciferous plants, but when the crop flowered they become more numerous there. When the crop ceased to flower new generation beetles moved to flowering verge plants.

6.0 Sugar Beet

Ingram (2000):

The report notes that as only the vegetative root of the plant is harvested, from plants that have not flowered, there is only an exceedingly small risk of contamination to the consumer through 'on-site' cross-pollination. Sugar beet are normally harvested before flowering and therefore the report gives no further recommendations about isolation distances. The extremely small risk emanates from the occasional flowering ('bolters').

NPRU Comment:

We support the view that contamination of the harvested sugar beet is unlikely for non-seed producing areas.

General comment on section 6

The report considers specifically cross pollination in non-seed areas but it is important to highlight the potential for growing sugar beet from contaminated seed. In such a case GM materials can be incorporated into the harvested product. Non-GM sugar beet can become cross pollinated with GM pollen in sugar beet seed producing areas. Subsequent seed from non-GM fields may then contain a proportion of GM/nonGM hybrid seed. A recent French study has shown that weed beet common in sugar beet producing areas can originate due to cross-pollination between 'wild' beet and cultivated beet in the sugar beet seed producing regions (Desplanque et al. 1999). The authors concluded that the transportation of crop/wild hybrids from the seed producing area to the sugar beet field is likely to be a recurrent phenomenon. Such studies (see also Boundry 1993) indicate the potential for GM/non-GM hybrid seed to be produced in seed producing areas and to be then grown in fields not intentionally GM.

Separation distances should be seen as one component of limiting transgene escape. Another important component that should also be considered in the case of sugar beet is the question of interfertile weed/wild relatives Studies by Vigouroux et al. (1999) and Raybould et al.( 1996;1998) have highlighted the potential for the introgression of sugar beet transgenes into wild beet populations.

References: