Following the introduction of dicamba-resistant (DR) soybean in 2017, concerns have increased with regard to dicamba off-target movement (OTM) onto sensitive crops, including vegetables. Field trials were conducted in New Jersey, New York, and Delaware to evaluate cucumber (‘Python’), eggplant (‘Santana’), and snap bean (‘Caprice’ and ‘Huntington’) injury and yield response to simulated dicamba drift rates. Crops were exposed to dicamba applied at 0, 0.056, 0.11, 0.56, 1.12, 2.24 g ae ha–1, representing 0, 1/10,000, 1/5,000, 1/1,000, 1/500, and 1/250 of the maximum soybean recommended label rate (560 g ae ha–1), respectively. Dicamba was applied either at the early vegetative (V2) or early reproductive (R1) stages. Minimal to no injury, vine growth reduction, or yield loss was noted for cucumber. Dicamba was more injurious to eggplant with up to 22% to 35% injury 2 wk after treatment (WAT) at rate ≥1.12 g ae ha–1; however, only the highest dicamba rate caused 27% reduction of the commercial yield compared to the nontreated control. Eggplant also showed greater sensitivity when dicamba exposure occurred at the R1 than at theV2 stage. Snap bean was the most sensitive crop investigated in this study. Injury 2 WAT was greater for ‘Caprice’ with dicamba ≥0.56 g ae ha–1 applied at V2 compared to R1 stage, whereas a similar difference occurred as low as 0.056 g ae ha–1 for ‘Huntington’. Compared to the nontreated control, reduction in plant height and biomass accumulation occurred for both cultivars at dicamba rate ≥0.56 g ae ha–1. Dicamba applied at 1.12 g ae ha–1 or greater resulted in 30% yield loss for ‘Caprice’, whereas ‘Huntington’ yield dropped 52% to 93% with dicamba ≥0.56 g ae ha–1. ‘Caprice’ bean yield was not influenced by dicamba timing of application. Conversely, ‘Huntington’ bean yield decreased by 8% following application at R1 compared to V2 stage.

A Palmer amaranth (Amaranthus palmeri S. Watson) population (KCTR: KS Conservation Tillage Resistant) collected from a conservation tillage field was confirmed with resistance to herbicides targeting at least six sites of action, including 2,4-D. The objectives of this research were using KCTR A. palmeri to investigate (1) the level of 2,4-D resistance, (2) 2,4-D absorption and translocation profiles, (3) the rate of 2,4-D metabolism compared with 2,4-D–tolerant wheat (Triticum aestivum L.), and (4) the possible role of cytochrome P450s (P450s) in mediating resistance. Dose–response experiments were conducted to assess the level of 2,4-D resistance in KCTR compared with susceptible plants, KSS (KS 2,4-D susceptible) and MSS (MS 2,4-D susceptible). KSS, MSS, and KCTR plants were treated with [14C]2,4-D to determine absorption, translocation, and metabolic patterns. Additionally, whole-plant dose–response assays were conducted by treating KCTR and KSS plants with P450 inhibitors (malathion, piperonyl butoxide [PBO]) before 2,4-D application. Dose–response experiments indicated a 6- to 11-fold 2,4-D resistance in KCTR compared with susceptible plants. No difference was found in percent [14C]2,4-D absorption among the populations. However, 10% less and 3 times slower translocation of [14C]2,4-D was found in KCTR compared with susceptible plants. Importantly, [14C]2,4-D was metabolized faster in KCTR than susceptible plants. At 24, 48, and 72 h after treatment (HAT), KCTR metabolized ∼20% to 30% more [14C]2,4-D than susceptible plants. KCTR plants and wheat generated metabolites with similar polarity. Nonetheless, at 24 HAT, ∼70% of [14C]2,4-D was metabolized in wheat, compared with only 30% in KCTR A. palmeri. Application of malathion before 2,4-D increased the sensitivity to 2,4-D in KCTR, suggesting involvement of P450s in mediating 2,4-D metabolism. However, no such impact of PBO was documented. Overall, this study confirms that enhanced metabolism is the primary mechanism of 2,4-D resistance in KCTR.

The developmental status of perennial weeds such as rush skeletonweed (Chondrilla juncea L.) can influence herbicide absorption and translocation. Differential efficacy between fall and spring applications suggests vernalization impacts herbicide absorption and translocation in other perennial asters. Clopyralid and aminopyralid absorption and translocation were quantified in nonvernalized and vernalized plants following application of 14C-labeled herbicides 2, 4, 8, 24, and 72 h after treatment. Less 14C clopyralid was absorbed, and at a slower rate, in vernalized plants. Movement out of the treated leaf was slower, with 14C clopyralid translocating more rapidly than 14C aminopyralid. More 14C moved to the roots in nonvernalized plants compared with vernalized plants, regardless of herbicide. Increased translocation to belowground survival structures is needed for effective control of C. juncea.

Off-target movement of dicamba and 2,4-D may injure and reduce the yield of many fruit and vegetable crops, impacting specialty crop producers and herbicide applicators alike. Two field experiments were established, using plant growth regulator–resistant soybean herbicide technologies, to evaluate drift and carryover risks to horseradish production. The drift experiment was conducted in 2015 and 2016 to evaluate impact of dicamba and 2,4-D simulated drift on horseradish production with a mid-POST application in soybean. Simulated drift rates were 1/10,000X, 1/1,000X, and 1/100X, with 1/2X, 1X, and 2X of standard application rates. Injury and yield loss was greater following application of 2,4-D than with dicamba. Yield reductions were observed beginning at the 1/1,000X rate of 2,4-D, with complete crop loss occurring when rates exceed 1/2X. In comparison, dicamba only reduced yields when applied at the 1X and 2X rates. Only horseradish roots from plants treated with dicamba at the 2X rate had greater dicamba residue than the nontreated control, and the amount detected, 0.32 parts per billion (ppb), was lower than the EPA tolerance of 100 ppb in root crops. There was little to no harvestable tissue for 2,4-D residue analysis for plants treated with 2,4-D at rates above 1/2X. The carryover experiment was a 2-yr rotational evaluation conducted in 2014, 2015, and 2016 to assess dicamba carryover to horseradish following application to dicamba-resistant soybean the previous season. Observations taken at 4, 6, and 8 wk after planting indicated no significant horseradish injury, nor was height, stand, or root weight reduced. These results suggest that horseradish growers should have few concerns about injury from dicamba drift or carryover. While 2,4-D applicators may need to be cautious when making applications near horseradish fields, 2,4-D may be an effective tool for controlling volunteer horseradish in 2,4-D–resistant soybean.

Research was conducted from 2015 to 2017 to investigate the potential for 2,4-D and multiple herbicide resistance in a waterhemp [Amaranthus tuberculatus (Moq.) J. D. Sauer] population from Missouri (designated MO-Ren). In the field, visual control of the MO-Ren population with 0.56 to 4.48 kg 2,4-D ha−1 ranged from 26% to 77% in 2015 and from 15% to 55% in 2016. The MO-Ren population was highly resistant to chlorimuron, with visual control never exceeding 7% either year. Estimates of the 2,4-D dose required to provide 50% visual control (I50) of the MO-Ren population were 1.44 kg ha−1 compared with only 0.47 kg 2,4-D ha−1 for the susceptible population. Based on comparisons to a susceptible population in dose–response experiments, the MO-Ren population was approximately 3-fold resistant to 2,4-D, and 7-, 7-, 22-, and 14-fold resistant to atrazine, fomesafen, glyphosate, and mesotrione, respectively. Dicamba and glufosinate were the only two herbicides that provided effective control of the MO-Ren population in these experiments. Examinations of multiple herbicide resistance at the individual plant level revealed that 16% of the plants of the MO-Ren population contained genes stacked for six-way herbicide resistance, and only 1% of plants were classified as resistant to a single herbicide (glyphosate). Results from these experiments confirm that the MO-Ren A. tuberculatus population is resistant to 2,4-D, atrazine, chlorimuron, fomesafen, glyphosate, and mesotrione, making this population the third 2,4-D–resistant A. tuberculatus population identified in the United States, and the first population resistant to six different herbicidal modes of action.

Aminocyclopyrachlor, a newly discovered synthetic auxin herbicide, and its methyl ester, appear to control a number of perennial broadleaf weeds. The potential volatility of this new herbicide and its methyl ester were determined under laboratory conditions and were also compared to dicamba and aminopyralid with the use of enclosed chamber and open-air plant bioassays. Bioassays consisting of visual estimates of epinastic responses and kidney bean and soybean leaf-width measurements were developed to measure vapor release from glass and plastic. Vapor release of aminocyclopyrachlor from glass surfaces was undetectable under laboratory conditions, and no phytotoxic responses were observed when plants were exposed to vapors emanating from various surfaces. Results were similar to those of aminopyralid, indicating the risk of plant injury from vapor movement of aminocyclopyrachlor and aminopyralid was very low. When combined with 1% methylated seed oil, vapor release of aminocyclopyrachlor-methyl ester reached 86% 192 h after application to glass surfaces. Phytotoxic responses of plants exposed to vapors emanating from various surfaces treated with aminocyclopyrachlor-methyl ester were similar to responses to dicamba under enclosed incubation conditions, but were less in outdoor, open-air environments. Studies are needed to understand better the risk of injury to nontarget plants due to vapor movement of aminocyclopyrachlor-methyl ester under field applications.

Two studies investigated off-target exposure of soybean to plant growth regulator (PGR) herbicides and determined if simultaneous exposure to PGR herbicides and labeled soybean herbicides increase PGR injury. The PGR herbicides, 2,4-D, clopyralid, and dicamba, as well as dicamba plus the auxin transport inhibitor diflufenzopyr, were applied to glyphosate-resistant soybean at the V3, V7, and R2 soybean growth stages. Two rates were chosen from previous and preliminary research to approximate threshold rates that would cause a yield reduction so as to distinguish differences in sensitivity between growth stages. All four PGR herbicides caused significant soybean injury, height reduction, and yield loss at one or more application rates and growth stages. Relative to other PGR herbicides, dicamba reduced soybean yield at the lowest rate (a potential rate from residues remaining in improperly cleaned application equipment), followed by clopyralid, with 2,4-D requiring the highest rate to reduce soybean yield (a potential rate from a high level of spray drift). Dicamba and dicamba plus diflufenzopyr were applied at equal fractions of labeled use rates for corn to compare them directly at equivalent levels of off-target movement. Dicamba plus diflufenzopyr caused less injury and yield loss than dicamba applied alone. In a second study, the highest labeled soybean use rates of glyphosate, imazethapyr, imazamox, and fomesafen were applied alone and in combination with the highest rate of dicamba used in the first study (1% of a labeled use rate for corn) at the V3 and V7 stages. Dicamba demonstrated synergistic interactions with imazamox, imazethapyr, and fomesafen (but not with glyphosate) to further reduce yield under some circumstances, especially when applied at the V7 stage. Several treatments that included dicamba reduced soybean seed weight when applied at either the V3 or V7 stage and reduced the number of seeds per pod at the V7 stage.

Field studies were conducted from 2003 through 2006 to evaluate bundleflower tolerance to several postemergence (POST) herbicides commonly used in pastures. All herbicides except imazapic, metsulfuron, and 2,4-DB injured bundleflower at least 88% when rated 4 wk after treatment (WAT). When rated 8 WAT, metsulfuron, picloram, and picloram plus 2,4-D injured bundleflower greater than 90%. Imazapic stunted bundleflower (10 to 35% injury) throughout the growing season. When rated 52 WAT, dicamba plus 2,4-D, picloram alone, and picloram plus 2,4-D injured bundleflower at least 97% in one of two years. Emergence of bundleflower with picloram-containing treatments ranged from 16 to 67% of the nontreated control, whereas emergence after dicamba-containing treatments ranged from 67 to 97%.

The primary goal of this research was to determine the inheritance of cross-resistance to several groups of auxinic herbicides through classical genetic approaches using auxinic herbicide–resistant (R) and –susceptible (S) wild mustard biotypes obtained from western Canada. F1 progeny were raised from crosses between homozygous auxinic herbicide–R and –S wild mustard parental lines. The F1 and F2 populations were assessed for picloram (pyridine group) and 2,4-D (phenoxyalkanoic group) resistance or susceptibility. Analyses of the F1 as well as the F2 progeny indicate that a single dominant gene confers the resistance to picloram and 2,4-D similar to an earlier report of dicamba-based (benzoic acid group) resistance in this wild mustard biotype. Furthermore, analyses of backcross progeny in this species indicate that resistance to all three auxinic herbicides, i.e., picloram, dicamba, and 2,4-D, is determined by closely linked genetic loci. With this information on inheritance of resistance to several auxinic herbicide families, the R biotype of wild mustard offers an excellent system to isolate and characterize the auxinic herbicide–resistance gene.

Auxin-mediated responses of kochia biotypes resistant to dicamba (HRd) or resistant to dicamba and fluroxypyr (HRdf) were compared with those of two susceptible biotypes. Rates of shoot and root gravitropic response and patterns of apical dominance, as determined by lateral bud sprouting after decapitation, were determined in the absence of herbicide treatment. Shoots of susceptible plants reoriented toward vertical at a rate of 23.4° h−1, whereas the rates of HRd and HRdf shoot reorientation were significantly slower at 7.2° and 14.4° h−1, respectively. Root gravitropic responses were not different between resistant and susceptible biotypes. In contrast to susceptible biotypes, both apical and basal lateral buds on HRd plants elongated after decapitation, although differences between HRd and susceptible biotypes became smaller during succeeding weeks. The elongation pattern of HRdf lateral buds was intermediate to that of susceptible and HRd plants. Inhibition assays of root growth by natural and synthetic auxins showed that HRd root growth was less sensitive to dicamba, 2,4-D, naphthalene-1-acetic acid, and indole-3-acetic acid than was root growth of HRdf or the susceptible biotypes. Collectively, results support the hypothesis that auxin binding or signal transduction pathways are impaired in resistant biotypes and that HRd may contain different lesions than does HRdf.

Aminopyralid is a new auxinic herbicide that provides Canada thistle control at lower use rates than clopyralid. Studies were conducted to determine if differences in absorption, translocation, or metabolism account for aminopyralid's greater biological activity. Radiolabeled aminopyralid and clopyralid were applied to individual leaves of rosette-stage Canada thistle plants. Nonionic surfactant was used for the absorption studies because it provided higher aminopyralid absorption than methylated seed oil or crop oil concentrate. Clopyralid was absorbed very rapidly, reaching 72% 24 h after treatment (HAT) and remaining near or above 80% during a 192-h time course. During the same time period, aminopyralid absorption increased from 34 to 60%. Clopyralid translocation out of the treated leaf was significantly higher than aminopyralid, 39% compared with 17%, respectively, 192 HAT. More of applied clopyralid translocated to aboveground tissue 192 HAT (27%) than to roots (12%), whereas aminopyralid translocation was similar in aboveground tissue (10%) and roots (7%) 192 HAT. Neither aminopyralid nor clopyralid was metabolized 192 HAT. Although aminopyralid is effective at lower use rates than clopyralid, clopyralid absorption and translocation were higher in Canada thistle. These results suggest that aminopyralid's chemical structure may provide for greater biological activity at the target site than clopyralid.

The broadleaf auxinic herbicide clopyralid was applied to three varieties of corn (Pioneer 36B08, Pioneer 3730, and Pioneer 3559) to determine whether it was phytotoxic to this crop. The effects of clopyralid on the growth and development of corn were compared with those induced by the auxinic herbicides dicamba, 2,4-D, picloram, and fluroxypyr. When compared with the other auxinic herbicides, clopyralid, applied as a foliar spray at the three- and six-leaf stages of development, caused the least damage to all three varieties of corn. Among the auxinic herbicides tested, fluroxypyr and dicamba caused severe damage to the three varieties, whereas picloram and 2,4-D had significant detrimental effects on the growth and development of varieties 36B08 and 3730. Similar results were also obtained when corn seeds were germinated in petri dishes containing increasing concentrations of the auxinic herbicides. In addition to correlating these growth and development effects with auxinic herbicide–induced physiological changes in corn, we compared the effects of clopyralid and dicamba on proton efflux from isolated protoplasts of the three varieties. Results of these biophysical studies are consistent with those from our growth and developmental studies and confirm that clopyralid is least effective in eliciting a response in corn when compared with dicamba. We conclude that clopyralid does not cause the deleterious effects seen with other auxinic herbicides when sprayed under optimal environmental conditions, i.e., high humidity and temperature.

Absorption and translocation of aminocyclopyrachlor (AMCP) was investigated in black walnut. Radiolabeled AMCP was applied foliarly as a free acid, and basally as a free-acid-formulated oil-soluble liquid and as a methyl ester emusifiable concentrate. Maximum absorption was 68% for the methyl ester, 47% for the free-acid oil-soluble liquid, and 8% for the free acid when evaluated as percent applied. When recovered AMCP was presented as Bq g−1, no difference was observed in AMCP absorption or translocation of 14C-labeled material out of treated plant parts. Recovery of applied radiolabeled material in the roots from basal applications was 5 to 11% 72 h after treatment (HAT), compared with 0.8% recovery from a foliar application. Regardless of chemical form or formulation type, comparable injury was observed within 72 HAT despite low levels of translocation.

The expression of a candidate auxin-responsive gene was evaluated for use in a diagnostic assay for plant growth regulator (PGR) herbicide injury in soybean leaves. Expression of GH3, a primary auxin-responsive gene, was evaluated in response to dicamba and clopyralid at the RNA and protein levels, and proteomic analysis evaluated global expression of proteins in response to dicamba. Expression of GH3 was also analyzed in response to heat, drought, salt stress, and infection by soybean mosaic virus (SMV) and bean pod mottle virus (BPMV) to determine the specificity of GH3 expression as a diagnostic marker for PGR herbicide injury. At the RNA level, GH3 was strongly induced by dicamba and clopyralid within 8 h after application. Expression peaked 1 to 3 d after treatment (DAT) in response to 10% and 1% of a labeled dose of dicamba and clopyralid, with higher expression levels detected at higher herbicide rates. At the protein level, GH3 expression was also strongly induced at 1, 2, and 3 DAT by 10% vs. 1% of a labeled dose of dicamba and clopyralid. Heat, drought, and salt stress and infection with SMV or BPMV had no effect on GH3 expression at either the RNA or protein level. Proteomic analysis identified three proteins that were up-regulated in response to dicamba. Two were induced for less than 7 DAT, and a third was identified as a stress-response enzyme (superoxide dismutase) that is likely not specific to PGR herbicide injury. Expression of GH3 was highly induced by PGR herbicides at the RNA and protein level and was not affected by environmental stresses or viral infection, indicating that GH3 expression has excellent potential for use in a diagnostic assay for PGR herbicide injury.

A waterhemp population from a native-grass seed production field in Nebraska was no longer effectively controlled by 2,4-D. Seed was collected from the site, and dose-response studies were conducted to determine if this population was herbicide resistant. In the greenhouse, plants from the putative resistant and a susceptible waterhemp population were treated with 0, 18, 35, 70, 140, 280, 560, 1,120, or 2,240 g ae ha−1 2,4-D. Visual injury estimates (I) were made 28 d after treatment (DAT), and plants were harvested and dry weights (GR) measured. The putative resistant population was approximately 10-fold more resistant to 2,4-D (R:S ratio) than the susceptible population based on both I50 (50% visual injury) and GR50 (50% reduction in dry weight) values. The R:S ratio increased to 19 and 111 as the data were extrapolated to I90 and GR90 estimates, respectively. GR50 doses of 995 g ha−1 for the resistant and 109 g ha−1 for the susceptible populations were estimated. A field dose-response study was conducted at the suspected resistant site with 2,4-D doses of 0, 140, 280, 560, 1,120, 2,240, 4,480, 8,960, 17,920, and 35,840 g ha−1. At 28 DAT, visual injury estimates were 44% in plots treated with 35,840 g ha−1. Some plants treated with the highest rate recovered and produced seed. Plants from the resistant and susceptible populations were also treated with 0, 9, 18, 35, 70, 140, 280, 560, or 1,120 g ae ha−1 dicamba in greenhouse bioassays. The 2,4-D resistant population was threefold less sensitive to dicamba based on I50 estimates but less than twofold less sensitive based on GR50 estimates. The synthetic auxins are the sixth mechanism-of-action herbicide group to which waterhemp has evolved resistance.

Auxinic herbicides are widely used for control of broadleaf weeds in cereal crops and turfgrass. These herbicides are structurally similar to the natural plant hormone auxin, and induce several of the same physiological and biochemical responses at low concentrations. After several decades of research to understand the auxin signal transduction pathway, the receptors for auxin binding and resultant biochemical and physiological responses have recently been discovered in plants. However, the precise mode of action for the auxinic herbicides is not completely understood despite their extensive use in agriculture for over six decades. Auxinic herbicide-resistant weed biotypes offer excellent model species for uncovering the mode of action as well as resistance to these compounds. Compared with other herbicide families, the incidence of resistance to auxinic herbicides is relatively low, with only 29 auxinic herbicide-resistant weed species discovered to date. The relatively low incidence of resistance to auxinic herbicides has been attributed to the presence of rare alleles imparting resistance in natural weed populations, the potential for fitness penalties due to mutations conferring resistance in weeds, and the complex mode of action of auxinic herbicides in sensitive dicot plants. This review discusses recent advances in the auxin signal transduction pathway and its relation to auxinic herbicide mode of action. Furthermore, comprehensive information about the genetics and inheritance of auxinic herbicide resistance and case studies examining mechanisms of resistance in auxinic herbicide-resistant broadleaf weed biotypes are provided. Within the context of recent findings pertaining to auxin biology and mechanisms of resistance to auxinic herbicides, agronomic implications of the evolution of resistance to these herbicides are discussed in light of new auxinic herbicide-resistant crops that will be commercialized in the near future.

Kochia plants resistant (R) to field rates of dicamba were characterized for their frequency of occurrence and levels of resistance and for the physiological fate of applied 14C-dicamba. Of 167 randomly sampled fields and seven fields identified by producers to contain R kochia, 19 contained plants that produced 1% or more R progeny. The maximum percentage of R progeny produced by parental plants from any field was 13%. An inbred R line derived from a field collection was 4.6-fold more resistant to dicamba than an inbred susceptible (S) line. Rates of 14C-dicamba uptake and translocation were similar in R and susceptible (S) plants up to 168 h after treatment (HAT). Concentrations of the primary metabolite, 5-hydroxy dicamba, were similar in R and S tissues up to 60 HAT, although amounts were significantly greater in R treated leaves by 96 and 168 HAT. However, because there were negligible levels of dicamba metabolites in R shoots and because the rate of metabolism was relatively slow, the observed changes were inadequate to account for observed resistance levels. Thus, dicamba resistance in kochia cannot be attributed to differential herbicide absorption, translocation, or metabolism. These findings, together with our field observations on the unusually slow spread of resistance within and among fields may indicate that dicamba resistance is a quantitative trait.