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Creeping b entgrass: when herbicide resistance goes wrong

Creeping b entgrass: when herbicide resistance goes wrong. Brigid Meints CROP 540 11/19/2012. Herbicide resistant weeds. Weeds become resistant to herbicides after repeated use; normal rates of the herbicide can no longer control the weed

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Creeping b entgrass: when herbicide resistance goes wrong

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  1. Creeping bentgrass: when herbicide resistance goes wrong Brigid Meints CROP 540 11/19/2012

  2. Herbicide resistant weeds • Weeds become resistant to herbicides after repeated use; normal rates of the herbicide can no longer control the weed • Transgenic plants engineered to be herbicide resistant that become weedy through gene flow

  3. Creeping Bentgrass (Agrostisstolonifera) • Specialty grass used for golf course greens, lawn bowling greens, and lawn tennis • High maintenance grass: requires high levels of fertilizer, frequent mowing, watering, aerating, and dethatching • Identification: bright green, fine textured, no auricles, long and tapered ligule • Perennial growth habit; spreads by stolons to form a mat or thatch layer above the soil line • Low-growing with a shallow root system • Allotetraploid • Obligate outcrosser: has very small seeds, but can also propagate asexually

  4. Background • Glyphosate-resistant creeping bentgrass created by Monsanto and Scotts: Event ASR368 • In January 2004, Monsanto and Scotts petitioned the Animal and Plant Health Inspection Service (APHIS) seeking a determination of nonregulated status for creeping bentgrass • APHIS began taking testimony to form an Environmental Impact Statement

  5. 14 species of Agrostis native to Oregon • Found in agronomic and nonagronomic habitats • Small seeds (6 million per pound) can be dispersed by wind, water, animals, or mechanical means • Naturally forms interspecific F1hybrids, • low in fertility or sterile • but under certain conditions have been shown to out-compete both parents • Interspecific hybrid: mating of two species, generally from the same genus. The offspring show a combination of traits and characteristics from the two parents. Offspring are often sterile. • Can hybridize with at least 12 other grass species from Agrostis and Polypogon • extensive clonal propagation is still possible

  6. Genetics of glyphosate resistance • Two components necessary: resistant target enzyme and sufficient expression of that enzyme within the transgenic plant • 5-Enol-pyruvylshikimate-3-phosphate synthase (EPSPS) is the target enzyme for the inhibition of glyphosate in the aromatic amino acid biosynthetic pathway • Agrobacterium sp. strain CP4 EPSPS was found to be an exceptional candidate. Does not have a negative impact on yield, quality, or nutritional value of the harvested crop • Inherited as a single Mendelianlocus

  7. History • 2003: • 162 ha of glyphosate-resistant transgenic creeping bentgrass tested under APHIS permit in central Oregon • Conducted within a 4453 ha control area established by the ODA • Located 144 km east of non-transgenic, commercial bentgrass production

  8. Watrudet al. (2004) • Majority of gene flow observed within 2 km (in direction of prominent winds) • Up to 21 km (sentinel plants) and 14 km (resident plants) • Found evidence of gene flow in 75 of 138 (sentinel) and 29 of 69 (resident) A. stolonifera plants based on seedling progeny survival after spraying in the greenhouse

  9. Sampling grid designed to test viable pollen flow from transgenic plants • Sentinel A. stolonifera plants cultivated in the Willamette Valley, transplanted in central Oregon (first tested for trait), transported very carefully

  10. PCR primers designed to test for the 1050-bp segment of the A. stolonifera CP4 EPSPS coding region. Matched GenBank accession for a CP4 EPSPS construct in glyphosate-resistant soybean

  11. Maximal distances of observed gene flow. Locations of sentinel A. stolonifera, resident A. stolonifera, and resident A. gigantea(A, B, and C respectively)

  12. Percent positive seedling progeny per location for sentinel A. stolonifera, resident A. stolonifera, and resident A. gigantea (A, B, and C respectively)

  13. Prevalence and incidence of plants positive for the transgene and seedling progeny

  14. Reichmanet al. (2006) • Resident populations of Agrostisspecies sampled in nonagronomic habitats outside of the ODA control zone • Attempted to discover parentage of plants positive for CP4 EPSPS, but Monsanto and Scotts had proprietary constraints • Used species-level molecular phylogenetic analyses

  15. Of 20,400 plant tissue samples taken, there were 9 positive plants (0.04%) • Distributed over 6 of the surveyed population areas, including the Crooked River National Grassland • Not possible to determine which field they came from, so distances are range • All near waterways or roadside • Gene trees showed that all wild transgenic plants had maternal and paternal A. stolonifera parents

  16. Why is this crop so problematic? • Size, density, and viability of the pollen • Wild, sexually compatible species • Floral synchrony between crop and wild species • Large source fields • Small seed size

  17. History • 2007: • APHIS completed their investigation into alleged compliance infractions by The Scotts Company. Scotts agreed to pay a civil penalty of $500,000, which is the maximum penalty allowed by the 2000 Plant Protection Act

  18. The saga continues… • 2012: • Reports of intergeneric hybridization with rabbitfootgrass, (Polypogonmonspeliensis) • Transgenesconfirmedusing PCR primers • Produced viable seed, had perennial growth habit and stolon production capability • Intergenerichybrid: hybrid between different genera

  19. Final thoughts • Careful consideration of potential gene flow when introducing transgenic crops • Consequences associated with that gene flow

  20. References • APHIS (2004) Monsanto Co. and The Scotts Co.; Availability of petition for determination of nonregulated status for genetically engineered glyphosate-tolerant creeping bentgrass. Federal Register 69, 315-317, January 5, 2004 • Heck et al. (2005). Development and Characterization of a CP4 EPSPS-Based, Glyphosate-Tolerant Corn Event:CropSci. 45:329-339 (2005). • Reichman, J. R., Watrud, l. S., Lee, E. H., Burdick, C. A., Bollman, M. A., Storm, M. J., King, G. A. And Mallory-Smith, C. (2006), Establishment of transgenic herbicide-resistant creeping bentgrass (Agrostisstolonifera L.) in nonagronomic habitats. Molecular Ecology, 15: 4243–4255. • Watrud LS, Lee EH, Fairbrother A et al. (2004) Evidence for landscape level pollen- mediated gene flow from genetically modified creeping bentgrass using CP4 EPSPS as a marker. Proceedings of the National Academy of Sciences, USA, 101, 14533–14538. • UC Davis. StatewideIntegratedPest Management: The UC Guide to HealthyLawns, CreepingBentgrass. http://www.ipm.ucdavis.edu/TOOLS/TURF/ TURFSPECIES creepbent.html • USDA. (2007). Release No. 0350.07. USDA concludes genetically engineered creeping bentgrass investigation. http://www.usda.gov/wps/portal/usda/ usdahome?contentidonly=true&contentid=2007/11/0350.xml • Zapiola, M. L. And Mallory-Smith, C. A. (2012), Crossing the divide: gene flow produces intergeneric hybrid in feral transgenic creeping bentgrass population. Molecular Ecology, 21: 4672–4680.

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