NOTE: The following articles refer to the "pyramid" strategy with transgenic toxins. Essentially this is the same as what's also referred to as gene stacking, ie where more than one transgene (in this case for pest resistance) is engineered into a plant.

It's been widely assumed till now that by having more than one transgene for pest resistance in a single plant, cross-resistance to the stacked pesticidal genes is made highly unlikely. Laboratory testing had also seemed to back this assumption up, and it's on this basis that regulatory authorities in N America recently allowed refuges of non-GM crops to be severely reduced where farmers were using the latest stacked gene products. The refuges required in corn (maize) have been reduced from 20% to just 5% and in cotton from 50% to 20%.
http://gmwatch.org/index.php?option=com_content&view=article&id=11317:smartstax-approval-maize-approvals-fail

All of this has been thrown into question by Tabashnik et al's unexpected finding that cross-resistance is possible. As the second item below, from the journal Nature,notes, "The results strike a cautionary note at a time when developers are racing to create crops that produce many different pesticides."

The net result is that stacked genes mean the risks for the consumer and the environment stack up in return for what could be unsustainable benefits for the farmer.
---
---
Pest Resistance to Pyramid Toxins is Possible
THIRD WORLD NETWORK BIOSAFETY INFORMATION SERVICE, 3 August 2009

Dear Friends and colleagues,

RE: Pest Resistance to Pyramid Toxins is Possible

To thwart pest resistance, some transgenic crops produce two different toxins targeting the same pest. This "pyramid" strategy is expected to work best when selection for resistance to one toxin does not cause cross-resistance to the other toxin. The most widely used pyramid is transgenic cotton producing Bacillus thuringiensis (Bt) toxins Cry1Ac and Cry2Ab, with which cross-resistance was presumed unlikely.

However, results show that cross-resistance between the two toxins is possible, suggesting that the pursuit of resistance-free transgenic crops may be impossible. The latest findings on resistance against Bt toxins in transgenic cotton are published in the Proceedings of the National Academy of Sciences.

The full text is available at: http://www.pnas.org/content/early/2009/07/02/0901351106.full.pdf+html?sid=ed940d 91-1b44-41c8-9dfe-> f51d6bf779d1.

With best wishes,
Third World Network
Email: This email address is being protected from spambots. You need JavaScript enabled to view it.
Website: www.biosafety-info.net and www.twnside.org.sg
---
---
1.Asymmetrical cross-resistance between Bacillus thuringiensis toxins Cry1Ac and Cry2Ab in pink bollworm
http://www.pnas.org/content/early/2009/07/02/0901351106.full.pdf+html?sid=ed940d 91-1b44-41c8-9dfe-%3E%20f51d6bf779d1

Full text:

Proceedings of the National Academy of Sciences, online publication

6 July 2009

Bruce E. Tabashnik (a, 1), Gopalan C. Unnithan (a), Luke Masson

(b), David W. Crowder (a), Xianchun Li (a), and Yves Carriere (a)

(a) Department of Entomology, University of Arizona, Tucson, AZ 85721; and
(b) Biotechnology Research Institute, National Research Council of Canada, Montreal, QC, Canada H4P 2R2

Edited by May R. Berenbaum, University of Illinois, Urbana, IL, and approved May 21, 2009 (received for review February 6, 2009)

Transgenic crops producing Bacillus thuringiensis (Bt) toxins kill some key insect pests and can reduce reliance on insecticide sprays. Sustainable use of such crops requires methods for delaying evolution of resistance by pests. To thwart pest resistance, some ransgenic crops produce 2 different Bt toxins targeting the same pest This "pyramid" strategy is expected to work best when selection for resistance to 1 toxin does not cause cross-resistance to the other toxin. The most widely used pyramid is transgenic cotton producing Bt toxins Cry1Ac and Cry2Ab. Cross-resistance between these toxins was presumed unlikely because they bind to different larval midgut target sites. Previous results showed that laboratory selection with Cry1Ac caused little or no cross-resistance to Cry2A toxins in pink bollworm (Pectinophora gossypiella), a major cotton pest. We show here, however, that laboratory selection of pink bollworm with Cry2Ab caused up to 420-fold cross-resistance to Cry1Ac as well as 240-fold resistance to Cry2Ab. Inheritance of resistance to high concentrations of Cry2Ab was recessive. Larvae from a laboratory strain resistant to Cry1Ac and Cry2Ab in diet bioassays survived on cotton bolls producing only Cry1Ac, but not on cotton bolls producing both toxins. Thus, the asymmetrical cross-resistance seen here does not threaten the efficacy of pyramided Bt cotton against pink bollworm. Nonetheless, the results here and previous evidence indicate that cross-resistance occurs between Cry1Ac and Cry2Ab in some key cotton ests. Incorporating the potential effects of such cross-resistance in resistance management plans may help to sustain the efficacy of pyramided Bt crops.
------------------------
Author contributions: B.E.T., GC.U., X.L., and Y.C. designed research; G.C.U. and L.M. performed research; B.E.T. and D.W.C. analyzed data; and B.E.T., G.C.U., L.M., D.W.C., X.L., and Y.C. wrote the paper.

Confict of interest statement: Although preparation of this article was not supported by organizations that may gain or lose financially through its publication, the authors have received support for other research from Monsanto, Cotton Inc., the Cotton Foundation, and the Arizona Cotton Growers Association. B.E.T. is a coauthor of a patent application filed with the World Intellectual Property Organization on engineering modified Bt toxins to counter pest resistance, which is related to published research [(2007) Science 318:1640 1642].

This article is a PNAS Direct Submission.

Freely available online through the PNAS open access option.

(1) To whom correspondence should be addressed. E-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.
---
---
2.Pests could overcome GM cotton toxins
Heidi Ledford
Nature, 6 July 2009, doi:10.1038
http://www.nature.com/news/2009/090706/full/news.2009.629.html

*Caterpillars reveal a chink in the armour of transgenic crops.

Laboratory studies suggest that it may be possible for insects to overcome two disparate toxins produced by genetically modified cotton. The results strike a cautionary note at a time when developers are racing to create crops that produce many different pesticides.

Insects can become resistant to individual insecticides in much the same way as bacteria develop resistance to antibiotics. One way to reduce this threat is to adopt a 'pyramid' approach and create crops that produce multiple toxins that target the same pest.

"This is the current trend of all the companies," says Juan Ferré, a geneticist at the University of Valencia in Spain. "They are all combining more than one gene to have better control and to delay resistance." For example, next year, Monsanto, a US agricultural products company based in St Louis, Missouri, intends to launch a line of maize (corn) that contains eight different genes that make the crop resistant to herbicides and to attack by insects.

One of the most common 'pyramided' crops on the market is cotton that produces two different 'Bt' toxins made naturally by the bacterium Bacillus thuringensis. The two toxic proteins, Cry1Ac and Cry2Ab, have very different amino-acid sequences and bind to different target sites.

As a result, mutations that confer resistance to both toxins were thought to be unlikely, says Bruce Tabashnik, an entomologist at the University of Arizona in Tucson. "The main way that insects become resistant is by altering the binding site of the toxin," he says. "These two toxins don't bind to the same site - if the insects altered the Cry1Ac binding site, it's not going to give cross resistance to Cry2Ab."

But when Tabashnik and his colleagues tried to selectively breed insects that were resistant to Cry2Ab, they found that that some were also resistant to Cry1Ac. The results are reported this week in Proceedings of the National Academy of Sciences (1).

Arms race

The researchers were studying pink bollworm (Pectinophora gossypiella) ”” a particular nuisance in the cotton fields of the southern United States. Crops expressing Cry1Ac have thus far largely held the pest at bay, and there has been no sign of Cry1Ac resistance emerging in the insects.

Tabashnik wanted to learn more about how insects may become resistant to the less-studied Cry2Ab protein, so the team raised a number of different laboratory strains of pink bollworms on a diet that contained the toxin. To their surprise, they generated a strain of pink bollworm that was not only resistant to 240-times higher levels of Cry2Ab than normal, but also to 420-times higher concentrations of Cry1Ac.

Although the binding sites of the two toxins differ, both toxins are activated via the same pathway in the insect. A change in the protease responsible for activating the toxins could provide an avenue to cross-resistance, Tabashnik says. Other changes in the insect's ability to cope with damaged cells could also play a part, says Ferré, who was not involved with the study.

The results show that cross-resistance between the two toxins is possible. But "this does not pose a threat for control by the current pyramided Bt cotton of this insect", Tabashnik says. The resistant pink bollworms were able to withstand high concentrations of both toxins in their diets, but they were not able to survive the higher concentrations of Cry2Ab found on cotton bolls produced by the pyramided transgenic cotton.

Ferré urges caution on extrapolating laboratory results to the field. "This is a special condition in the laboratory," he says. "The important thing is to find out whether that resistance can be obtained in the field."

Nevertheless, the results do highlight the continued threat of resistance, adds Tabashnik. "Pyramids are not a panacea," he says. "Evolution by insects is not something that scientists are going to stop."

References

1. Tabashnik, B. E. et al. Proc. Natl Acad. Sci. USA advance online publication doi:10.1073/pnas.0901351106 (2009).