Gene amplification confers glyphosate resistance in Amaranthus palmeri

The herbicide glyphosate became widely used in the United States and other parts of the world after the commercialization of glyphosate-resistant crops. These crops have constitutive overexpression of a glyphosate-insensitive form of the herbicide target site gene, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). Increased use of glyphosate over multiple years imposes selective genetic pressure on weed populations. We investigated recently discovered glyphosate-resistant Amaranthus palmeri populations from Georgia, in comparison with normally sensitive populations. EPSPS enzyme activity from resistant and susceptible plants was equally inhibited by glyphosate, which led us to use quantitative PCR to measure relative copy numbers of the EPSPS gene. Genomes of resistant plants contained from 5-fold to more than 160-fold more copies of the EPSPS gene than did genomes of susceptible plants. Quantitative RT-PCR on cDNA revealed that EPSPS expression was positively correlated with genomic EPSPS relative copy number. Immunoblot analyses showed that increased EPSPS protein level also correlated with EPSPS genomic copy number. EPSPS gene amplification was heritable, correlated with resistance in pseudo-F2 populations, and is proposed to be the molecular basis of glyphosate resistance. FISH revealed that EPSPS genes were present on every chromosome and, therefore, gene amplification was likely not caused by unequal chromosome crossing over. This occurrence of gene amplification as an herbicide resistance mechanism in a naturally occurring weed population is particularly significant because it could threaten the sustainable use of glyphosate-resistant crop technology

Bioensaios para identificação de biótipos de Euphorbia heterophylla com resistência múltipla a inibidores da ALS e da PROTOX Greenhouse and laboratory bioassays for identification of Euphorbia heterophylla biotypes with multiple resistance to PROTOX and ALS-inhibiting herbicides

Quatro bioensaios, dois em casa de vegetação e dois em laboratório, foram conduzidos com o objetivo de identificar biótipos de Euphorbia heterophylla (EPHHL) com resistência múltipla a inibidores da ALS e da PROTOX. Em casa de vegetação, plantas do biótipo suscetível (S) e dos biótipos 4 e 23, suspeitos de resistência múltipla, foram aspergidas com diferentes doses de imazethapyr e fomesafen. Nos bioensaios em laboratório, sementes dos biótipos S e 4 foram depositadas em placas de Petri contendo diferentes concentrações dos mesmos herbicidas. Curvas de dose-resposta foram ajustadas, utilizando os modelos logístico e polinomial, respectivamente, para os dados obtidos em casa de vegetação e em laboratório. Em casa de vegetação, o fator de resistência (FR) a imazethapyr para o biótipo 4 foi superior a 24, enquanto para o biótipo 23 ele foi de 15. Os FRs a fomesafen foram, respectivamente, de 62 e 39, para os mesmos biótipos. Em um período de 144 horas, concentrações de imazethapyr e fomesafen no bioensaio em laboratório foram capazes de discriminar os crescimentos da parte aérea e radicular dos biótipos de EPHHL com resistência múltipla e S. Os resultados confirmam ser tanto os testes em casa de vegetação quanto os laboratoriais, utilizando placas de Petri, metodologias apropriadas para discriminar biótipos de EPHHL S daqueles com resistência múltipla.<br>Four bioassays were developed under greenhouse and laboratory conditions to identify Euphorbia heterophylla (EPHHL) biotypes with multiple resistance to PROTOX and ALS-inhibiting herbicides. In the greenhouse bioassays, plants of a susceptible (S) biotype and of two biotypes suspected of multiple resistance (#4 and #23) were sprayed using different levels of the herbicides imazethapyr and fomesafen. The laboratory bioassays consisted of a comparative test between biotypes S and #4, exposed to germination in imazethapyr and fomesafen solutions. Dose-response relationships were adjusted using logistic and polynomial models, respectively, for data obtained from greenhouse and laboratory bioassays. For biotype #4, the resistance factor (RF) to imazethapyr was above 24, while for biotype #23 it was 15. FR to fomesafen were, respectively, 62 and 39, for the same biotypes. In the laboratory bioassay, a period of 144 hours was sufficient to discriminate differencial growth response to imazethapyr and fomesafen levels between R and S biotypes. The results confirm that both greenhouse and laboratory tests with Petri dishes are appropriate methodologies to discriminate S and multiple R EPHHL biotypes