Transcriptomics and population genetics of glyphosate-resistant Conyza bonariensis (fleabane) populations.

Abstract

Glyphosate-resistant populations of Conyza bonariensis were first detected in 2005 in northeastern cropping regions of Australia. The EPSPS gene is present in three copies in this species: EPSPS1 and EPSPS2 are the result of an ancient duplication, but EPSPS2 and EPSPS3 diverged more recently. Initial Sanger sequencing independently targeting EPSPS1 and EPSPS2 indicated no target-site mutations, and a transcriptomics approach was taken to investigate non-target-site resistance. Three individuals from each of five lines (two susceptible and three resistant) were sampled for RNAseq prior to, and 48 hours after, glyphosate treatment. Many genes were differentially expressed (DE) following glyphosate treatment (over 20,000 in all lines) but more genes were DE in susceptible lines than resistant lines. A list of all DE genes in susceptible lines was constructed, representing the differential expression of genes as part of the glyphosate-induced death process. When these genes were subtracted from the list of DE genes in resistant lines, under 4000 DE genes remained. One of these (a member of the major facilitator superfamily of genes) is involved in cell transport and is a candidate for non-targetsite resistance. There were no differences in EPSPS gene expression across susceptible and resistant lines. When the transcriptome reads were mapped to each of the three EPSPS genes independently, however, there was a low level of target-site mutation (2 to 10% of reads) in EPSPS2. Because C. bonariensis is an allopolyploid with three genome progenitors, each EPSPS gene is present in six copies, and the number of target-site resistant alleles detected is equivalent to one copy out of six for the EPSPS3 gene. This low level of target-site resistance does not play a major role in resistance because it is present both in very strong and very weak resistant lines.