Investigation of glyphosate resistance in Phragmites australis

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Jordan Nikkel, Rachel Pash, Young D. Choi and Radmila Stanic, Biological Sciences, Purdue University Northwest, Hammond, IN

Background/Question/Methods
Common reed (Phragmites australis) is a highly invasive plant species with cosmopolitan distribution that has posed itself as a major ecological problem across the globe. Phragmites invades new areas rapidly and forms dense, monospecific stands which greatly reduce plant diversity. Management strategies for this species in many areas have focused on repeated applications of the herbicide glyphosate. These practices have had some success over short periods of time, but the long-term effectiveness remains in doubt as reinvasion often occurs and some populations seem to be unaffected by treatment.
Glyphosate operates by interfering with the enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), which plays a role in the Shikimic acid pathway. Resistance to glyphosate has been documented in a number of plant species. One mechanism is the result of a pair of point mutations within the region of DNA coding for EPSPS. This mutation is referred to as TIPS, as it results from the changing of a threonine to an isoleucine and a proline to a serine at positions 102 and 106 of the amino acid chain, respectively. Given the frequent application of glyphosate to populations of Phragmites, it is possible that this species could also be developing resistance.
We investigated potential TIPS mutations in the region of DNA that codes for EPSPS in Phragmites. Foliar tissue samples were frozen at -80 °C after being collected from three clumps of Phragmites within the Indiana Dunes National Park. The first clump was never sprayed with glyphosate, the second was sprayed but recovered to robust growth, and the third has not completely recovered from recent spraying. DNA was isolated and amplified using primers specific for the target region. Samples that displayed strong bands during agarose gel electrophoresis were sent away for sequencing for later analysis.

Results/Conclusions
We found mutations in the target codons in two samples (n=22). In one from the robust clump, a codon ACT (prescribe threonine) was replaced by GCT (alanine). The mutation in the other sample did not result in an amino acid change. However, the results were deemed inconclusive because of a lack of replicates in other samples and incomplete sequencing around the targeted codons. Future studies may expand upon our work by looking for these mutations in other populations or attempting to explore other resistance mechanisms. Evidence for glyphosate resistance in Phragmites would have major implications for the management of this species and should be investigated wherever glyphosate is used.