During the past 30 yr an impasse has developed in the discovery and commercialization of synthetic herbicides with new molecular targets and novel chemistries. Similarly, there has been little success with bioherbicides, both microbial and chemical. These bioherbicides are needed to combat fast-growing herbicide resistance and to fulfill the need for more environmentally and toxicologically safe herbicides. In response to this substantial and growing opportunity, numerous start-up companies are utilizing novel approaches to provide new tools for weed management. These diverse new tools broaden the scope of discovery, encompassing advanced computational, bioinformatic, and imaging platforms; plant genome–editing and targeted protein degradation technologies; and machine learning and artificial intelligence (AI)-based strategies. This review contains summaries of the presentations of 10 such companies that took part in a symposium held at the WSSA annual meeting in 2024. Four of the companies are developing microbial bioherbicides or natural product–based herbicides, and the other six are using advanced technologies, such as AI, to accelerate the discovery of herbicides with novel molecular target sites or to develop non-GMO, herbicide-resistant crops.

Eurasian watermilfoil (Myriophyllum spicatum L.) is an invasive aquatic plant that can hybridize with the native northern watermilfoil (M. sibiricum Kom.). These milfoil hybrids (M. spicatum × M. sibiricum) are becoming more prevalent in many lakes where the invasive and the native milfoil co-occur. Hybrid plants are more vigorous than either parent with a faster growth rate and lower sensitivity to some herbicides. The aquatic herbicides endothall and 2,4-D provide two effective modes of action for management of the hybrids. For more than a decade, these two herbicides have been used in combination as an effective control option and a resistance management strategy. How this combination impacts herbicide movement and efficacy is unknown. Therefore, the objective of this research was to determine the activity of endothall and 2,4-D combined compared with activity applied alone. Absorption and translocation of endothall, 2,4-D, and the combination was determined in hybrid plants over a 96-h time course. Endothall accumulation was not impacted when these herbicides were applied in combination; however, 2,4-D accumulation increased by 80%, relative to when 2,4-D was applied alone. Endothall translocation from shoots to roots decreased by almost 50% when applied in combination with 2,4-D (alone = 16.7% ± 2.6%; combination = 9.2% ± 1.2%). Shoot-to-root translocation of 2,4-D also decreased when the two herbicides were applied in combination (24.8% ± 2.6% when applied alone to only 3.93% ± 0.4% when in the presence of endothall). This research demonstrates that combining herbicides can significantly impact herbicide activity in plants. Future research is needed to determine whether this reduced translocation negatively impacts operational effectiveness when these herbicides are applied in combination.

Due to increased food demand, the use of herbicides is both necessary and on the rise. Several herbicide classes target photosynthetic electron transport: Herbicide Resistance Action Committee (HRAC) Groups 5, 6, and 22. These herbicides are used in large amounts in many different cropping systems to control several species of broadleaf and grass weeds. This article provides a comprehensive review of what these photosynthesis inhibitors are, how they are used, and their modes of action. Presently, commercial herbicides only inhibit electron flow at two different sites: photosystem II (PSII) and photosystem I (PSI). Herbicides that inhibit electron flow at PSII block the movement of electrons down the electron transport chain, while those that inhibit electron flow at PSI accept electrons. Necrosis developing on the leaves of plants treated with PSII and PSI inhibitors is due to the accumulation of reactive oxygen species. Evolution of resistance, toxicity concerns, and other limitations of these herbicides call for the exploration of new chemistries that can be used to target this pathway.

Soil-applied herbicides are important for controlling weeds in many crops but risk damage to susceptible rotational crops if they persist. Field studies were conducted in Powell, WY, from 2015 through 2017 to evaluate the effect of reduced water availability on soil-applied herbicide dissipation. Eight soil-applied herbicides, applied to dry bean or corn, were exposed to three season-long irrigation treatments (100%, 85%, and 70% of estimated crop evapotranspiration [ETc]) by overhead sprinkler. Soil samples were collected to a depth of 10 cm from 0 to 140 d after application, and soil herbicide concentrations were quantified using gas or liquid chromatography and mass spectrometry. Herbicide concentrations were regressed over time to produce a soil half-life estimate for each herbicide and irrigation treatment. Reduced irrigation decreased dry bean yield by up to 77% and corn yield by up to 50%. After adjusting for precipitation, the lowest irrigation treatment received 78% and 76% as much water as the full irrigation treatment in 2015 and 2016, respectively. This significantly increased the soil half-life of imazethapyr but did not increase the soil half-life of atrazine, pyroxasulfone, saflufenacil, ethalfluralin, trifluralin, or pendimethalin. Reduced irrigation did not increase carryover injury to rotational crops from these herbicides 1 yr after application. Instead, carryover response was determined by the inherent persistence of individual herbicides. Imazethapyr (0.1 kg ai ha−1) injured rotational sugar beet, and isoxaflutole (0.1 kg ai ha−1) injured rotational dry bean. Pyroxasulfone (0.2 kg ai ha−1), atrazine (2.0 kg ai ha−1), saflufenacil (0.1 kg ai ha−1) + dimethenamid-P (0.6 kg ai ha−1), ethalfluralin (0.8 kg ai ha−1), trifluralin (0.6 kg ai ha−1), and pendimethalin (1.1 kg ai ha−1) did not injure rotational crops regardless of irrigation treatment. Drought stress sufficient to cause up to 77% crop yield loss did not increase soil-applied herbicide carryover.

Protoporphyrinogen oxidase (PPO)-inhibiting herbicides remain an important and useful chemistry 60 yr after their first introduction. In this review, based on topics introduced at the Weed Science Society of America 2021 symposium titled “A History, Overview, and Plan of Action on PPO Inhibiting Herbicides,” we discuss the current state of PPO-inhibiting herbicides. Renewed interest in the PPO-inhibiting herbicides in recent years, due to increased use and increased cases of resistance, has led to refinements in knowledge regarding the mechanism of action of PPO inhibitors. Herein we discuss the importance of the two isoforms of PPO in plants, compile a current knowledge of target-site resistance mechanisms, examine non–target site resistance cases, and review crop selectivity mechanisms. Consistent and reproducible greenhouse screening and target-site mutation assays are necessary to effectively study and compare PPO-inhibitor resistance cases. To this end, we cover best practices in screening to accurately identify resistance ratios and properly interpret common screens for point mutations. The future of effective and sustainable PPO-inhibitor use relies on development of new chemistries that maintain activity on resistant biotypes and the promotion of responsible stewardship of PPO inhibitors both new and old. We present the biorational design of the new PPO inhibitor trifludimoxazin to highlight the future of PPO-inhibitor development and discuss the elements of sustainable weed control programs using PPO inhibitors, as well as how responsible stewardship can be incentivized. The sustained use of PPO inhibitors in future agriculture relies on the effective and timely communication from mode of action and resistance research to agronomists, Extension workers, and farmers.

The herbicides that inhibit 4-hydroxyphenylpyruvate dioxygenase (HPPD) are primarily used for weed control in corn, barley, oat, rice, sorghum, sugarcane, and wheat production fields in the United States. The objectives of this review were to summarize 1) the history of HPPD-inhibitor herbicides and their use in the United States; 2) HPPD-inhibitor resistant weeds, their mechanism of resistance, and management; 3) interaction of HPPD-inhibitor herbicides with other herbicides; and 4) the future of HPPD-inhibitor-resistant crops. As of 2022, three broadleaf weeds (Palmer amaranth, waterhemp, and wild radish) have evolved resistance to the HPPD inhibitor. The predominance of metabolic resistance to HPPD inhibitor was found in aforementioned three weed species. Management of HPPD-inhibitor-resistant weeds can be accomplished using alternate herbicides such as glyphosate, glufosinate, 2,4-D, or dicamba; however, metabolic resistance poses a serious challenge, because the weeds may be cross-resistant to other herbicide sites of action, leading to limited herbicide options. An HPPD-inhibitor herbicide is commonly applied with a photosystem II (PS II) inhibitor to increase efficacy and weed control spectrum. The synergism with an HPPD inhibitor arises from depletion of plastoquinones, which allows increased binding of a PS II inhibitor to the D1 protein. New HPPD inhibitors from the azole carboxamides class are in development and expected to be available in the near future. HPPD-inhibitor-resistant crops have been developed through overexpression of a resistant bacterial HPPD enzyme in plants and the overexpression of transgenes for HPPD and a microbial gene that enhances the production of the HPPD substrate. Isoxaflutole-resistant soybean is commercially available, and it is expected that soybean resistant to other HPPD inhibitor herbicides such as mesotrione, stacked with resistance to other herbicides, will be available in the near future.

Hydrilla [Hydrilla verticillata (L. f.) Royle] is often called the “perfect aquatic weed,” as it has numerous physiological adaptations that make it highly aggressive and competitive. Hydrilla verticillata has historically been managed effectively using fluridone; however, the overreliance on this single mechanism of action (MOA) resulted in evolved fluridone resistance in the late 1990s. Where fluridone-resistant H. verticillata populations evolved, endothall became widely used for H. verticillata control. In 2018, florpyrauxifen-benzyl, a highly active auxin-mimic herbicide, was registered for H. verticillata control, and its use has increased since its introduction. Endothall and florpyrauxifen-benzyl provide two effective MOAs for H. verticillata management, and combining these two MOAs would be an effective strategy to delay further resistance evolution. The objective of this research was to determine whether combining endothall and florpyrauxifen-benzyl would significantly impact the behavior of either herbicide in dioecious (DHV) or monoecious (MHV) H. verticillata compared with their behavior when applied alone. Endothall and florpyrauxifen-benzyl absorption and accumulation alone and in combination were measured over a 192-h time course. Translocation patterns were also determined. Herbicide accumulation in MHV and DHV was not impacted when these herbicides were applied in combination. Endothall translocation from shoots to roots in DHV was not impacted (alone = 18.7 ± 1.4%; combination = 23.2 ± 2.2%); however, endothall shoot-to-root translocation in MHV was reduced from 16.2 ± 1.3% applied alone to 2.2 ± 0.1% when applied in combination with florpyrauxifen-benzyl. Florpyrauxifen-benzyl shoot-to-root translocation was reduced in both MHV and DHV when applied in combination with endothall. Florpyrauxifen-benzyl translocation was reduced by 16- and 6-fold in DHV and MHV, respectively. These data do not suggest that there would be operational impacts from endothall and florpyrauxifen-benzyl mixtures. Still, there appear to be changes in herbicide behavior, primarily shoot-to-root translocation, when these two herbicides are applied in combination.

Wild poinsettia (Euphorbia heterophylla L.) is a troublesome broadleaf weed in grain production areas in South America. Herbicide resistance to multiple sites of action has been documented in this species, including protoporphyrinogen oxidase (PPO) inhibitors. We investigated the physiological and molecular bases for PPO-inhibitor resistance in a E. heterophylla population (RPPO) from Southern Brazil. Whole-plant dose–response experiments revealed a cross-resistance profile to three different chemical groups of PPO inhibitors. Based on dose–response parameters, RPPO was resistant to lactofen (47.7-fold), saflufenacil (8.6-fold), and pyraflufen-ethyl (3.5-fold). Twenty-four hours after lactofen treatment (120 g ha−1) POST, RPPO accumulated 27 times less protoporphyrin than the susceptible population (SPPO). In addition, RPPO generated 5 and 4.5 times less hydrogen peroxide and superoxide than SPPO, respectively. The chloroplast PPO (PPO1) sequences were identical between the two populations, whereas 35 single-nucleotide polymorphisms were found for the mitochondrial PPO (PPO2). Based on protein homology modeling, the Arg-128-Leu (homologous to Arg-98-Leu in common ragweed [Ambrosia artemisiifolia L.] was the only one located near the catalytic site, also in a conserved region of PPO2. The cytochrome P450 monooxygenase inhibitor malathion did not reverse resistance to lactofen in RPPO, and both populations showed similar levels of PPO1 and PPO2 expression, suggesting that metabolic resistance and PPO overexpression are unlikely. This is the first report of an Arg-128-Leu mutation in PPO2 conferring cross-resistance to PPO inhibitors in E. heterophylla.

Glufosinate inhibits glutamine synthetase (GS), a key enzyme for amino acid metabolism and photorespiration. Protoporphyrinogen oxidase (PPO) inhibitors block chlorophyll biosynthesis and cause protoporphyrin accumulation, a highly photodynamic intermediate. Both herbicides ultimately lead to plant death by a massive accumulation of reactive oxygen species (ROS) through different mechanisms. We investigated a potential synergistic effect by the mixture of the two herbicide mechanisms of action (MoAs). The tank mix between a low rate of glufosinate (280 g ai ha−1) with an ultra-low dose of saflufenacil (1 g ha−1) provided enhanced herbicidal activity compared with the products applied individually on Palmer amaranth (Amaranthus palmeri S. Watson). The synergism between the two herbicides was also confirmed by isobole analysis and field trials. The herbicide combination provided high levels of efficacy when applied at low temperature and low humidity. Mechanistically, glufosinate caused a transient accumulation of glutamate, the building block for chlorophyll biosynthesis. Consequently, inhibition of both GS and PPO resulted in greater accumulation of protoporphyrin and ROS, forming the physiological basis for the synergism between glufosinate and PPO inhibitors. While the synergy between the two herbicide MoAs provided excellent efficacy on weeds, it caused low injury to PPO-resistant waterhemp [Amaranthus tuberculatus (Moq.) Sauer] and high injury to both glufosinate-resistant and glufosinate-susceptible soybean [Glycine max (L.) Merr.]. Glufosinate enhances the activity of PPO inhibitors through glutamate and protoporphyrin accumulation, leading to increased levels of ROS and lipid peroxidation. The synergism between the two herbicide MoAs can help to overcome environmental effects limiting the efficacy of glufosinate. Future research is needed to optimize the uses for this herbicidal composition across different cropping systems.

Invasive winter annual grass infestations on rangeland accumulate large quantities of litter on the soil surface, as plants senesce yearly and decompose slowly. It has been speculated that winter annual grass litter can adsorb soil-active herbicides and reduce overall performance. Three experiments were conducted from 2017 to 2018 at the Colorado State University Weed Research Laboratory to evaluate interception and subsequent desorption of herbicides applied to litter from three invasive winter annual grass species with simulated rainfall. Imazapic, rimsulfuron, and indaziflam were applied to medusahead [Taeniatherum caput-medusae (L.) Nevski], ventenata [Ventenata dubia (Leers) Coss.], and downy brome (Bromus tectorum L.) litter at two amounts (equivalent to 1,300 and 2,600 kg ha−1). Rainfall was simulated at 3, 6, 12, and 24 mm at 0, 1, and 7 d after herbicide application. Herbicide concentration from the collected rainfall was measured using liquid chromatography–tandem mass spectrometry. At 2,600 kg ha−1, B. tectorum herbicide interception was 84.3%, while V. dubia and T. caput-medusae averaged 76% herbicide interception. There were no differences in desorption among the three litter types. Simulated rainfall at 0 d after application recovered 100% of the intercepted rimsulfuron and imazapic from B. tectorum litter, while recovery decreased to 65% with rainfall at 1 or 7 d after application. Only 54% of indaziflam could be recovered at 0 d, and recovery decreased to 33% when rainfall was applied at 1 or 7 d after application. Applying soil-active herbicides before forecasted rain or tank mixing with a POST herbicide to provide initial control could potentially increase the amount of herbicide reaching the soil and provide more consistent invasive winter annual grass control.

Several grass and broadleaf weed species around the world have evolved multiple-herbicide resistance at alarmingly increasing rates. Research on the biochemical and molecular resistance mechanisms of multiple-resistant weed populations indicate a prevalence of herbicide metabolism catalyzed by enzyme systems such as cytochrome P450 monooxygenases and glutathione S-transferases and, to a lesser extent, by glucosyl transferases. A symposium was conducted to gain an understanding of the current state of research on metabolic resistance mechanisms in weed species that pose major management problems around the world. These topics, as well as future directions of investigations that were identified in the symposium, are summarized herein. In addition, the latest information on selected topics such as the role of safeners in inducing crop tolerance to herbicides, selectivity to clomazone, glyphosate metabolism in crops and weeds, and bioactivation of natural molecules is reviewed.

Glyphosate-resistant (GR) goosegrass [Eleusine indica (L.) Gaertn.] was recently identified in Brazil, but its resistance mechanism was unknown. This study elucidated the resistance mechanism in this species and developed a molecular marker for rapid detection of this target-site resistance trait. The resistance factor for the resistant biotype was 4.4-fold compared with the glyphosate-susceptible (GS) in greenhouse dose–response experiments. This was accompanied by a similar (4-fold) difference in the levels of in vitro and in planta shikimate accumulation in these biotypes. However, there was no difference in uptake, translocation, or metabolism of glyphosate between the GS and GR biotypes. Moreover, both biotypes showed similar values for 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) copy number and transcription. Sequencing of a 330-bp fragment of the EPSPS gene identified a single-nucleotide polymorphism that led to a Pro-106-Ser amino acid substitution in the enzyme from the GR biotype. This mutation imparted a 3.8-fold increase in the amount of glyphosate required to inhibit 50% of EPSPS activity, confirming the role of this amino acid substitution in resistance to glyphosate. A quantitative PCR–based genotyping assay was developed for the rapid detection of resistant plants containing this Pro-106-Ser mutation.

In recent years, herbicide resistance has attracted much attention as an increasingly urgent problem worldwide. Unfortunately, most of that effort was focused on confirmation of resistance and characterization of the mechanisms of resistance. For management purposes, knowledge about biology and ecology of the resistant weed phenotypes is critical. This includes fitness of the resistant biotypes compared with the corresponding wild biotypes. Accordingly, fitness has been the subject of many studies; however, lack of consensus on the concept of fitness resulted in poor experimental designs and misinterpretation of the ensuing data. In recent years, methodological protocols for conducting proper fitness studies have been proposed; however, we think these methods should be reconsidered from a herbicide-resistance management viewpoint. In addition, a discussion of the inherent challenges associated with fitness cost studies is pertinent. We believe that the methodological requirements for fitness studies of herbicide-resistant weed biotypes might differ from those applied in other scientific disciplines such as evolutionary ecology and genetics. Moreover, another important question is to what extent controlling genetic background is necessary when the aim of a fitness study is developing management practices for resistant biotypes. Among the methods available to control genetic background, we suggest two approaches (single population and pedigreed lines) as the most appropriate methods to detect differences between resistant (R) and susceptible (S) populations and to derive herbicide-resistant weed management programs. Based on these two methods, we suggest two new approaches that we named the “recurrent single population” and “recurrent pedigreed lines” methods. Importantly, whenever the aim of a fitness study is to develop optimal resistance management, we suggest selecting R and S plants within a single population and evaluating all fitness components from seed to seed instead of measuring changes in the frequency of R and S alleles through multigenerational fitness studies.

Greenhouse-grown soybean cultivars varied in their tolerance to preemergence application of sulfentrazone. The cultivars Ransom, Hutcheson, Kato, Gasoy 17, and Cobb exhibited relatively low tolerance to 0.5 kg ai ha−1 sulfentrazone with 38, 41, 46, 50, and 58% height reduction compared to respective controls. The growth of tolerant cultivars Centennial, Edison, and Hartz 5164 was not affected by this treatment. However, the growth of all cultivars was reduced at the excessive rate of 2.0 kg ha−1 preemergence application of sulfentrazone. No differences in root uptake or translocation of [14C] sulfentrazone were observed between the relatively tolerant and less tolerant cultivars tested. Centennial and Hutcheson cultivars rapidly metabolized sulfentrazone via oxidative degradation of the 3-methyl group on the triazolinone ring of the herbicide. Only 4.7 and 4.9% of the active ingredient remained in the foliage of Hutcheson and Centennial 24 h after treatment, respectively. While there were no differences in Protox inhibition or Proto IX accumulation between the two cultivars, Hutcheson was more sensitive than Centennial to peroxidative stresses induced by either Proto IX accumulation or rose bengal. Therefore, tolerance to sulfentrazone is due to rapid metabolism of the herbicide; however, the intraspecific difference in response to sulfentrazone appears to be due to intrinsic differential tolerance to the herbicide-induced peroxidative stress.

Consistent with field observations, sicklepod exhibited considerable tolerance to sulfentrazone, and coffee senna showed relatively high sensitivity to this herbicide in greenhouse tests. Germination was not inhibited in either species at up to 12.9 μM of the herbicide. However, the chlorophyll content of herbicide-treated coffee senna cotyledonary leaves was greatly reduced, and seedlings died within 10 d after treatment, while sicklepod seedlings were not visibly affected. Shoot height of coffee senna was inhibited 90% by sulfentrazone at 0.5 kg ai ha−1, while the growth of sicklepod was not affected up to 2.0 kg ai ha−1. Root uptake of radiolabeled sulfentrazone was 74% greater in coffee senna than sicklepod, but the amount of radioactivity recovered from the shoots of both species after 12 h was not different. Eighty-three percent of the parent compound remained in coffee senna leaf tissue after 9 h root exposure to the herbicide. In contrast, sicklepod took up relatively less sulfentrazone through the root and metabolized sulfentrazone in the foliage more rapidly than coffee senna, with 91.6% of the herbicide being metabolized during the first 9 h of exposure. These results suggest that the tolerance of sicklepod to sulfentrazone is primarily due to a relatively high rate of metabolism of the herbicide compared to coffee senna.

Sulfentrazone was foliar applied at 34 and 56 g ai ha−1 alone or in combination with surfactants to soybean cultivars Hutcheson and Centennial and to sicklepod, coffee senna, smallflower morningglory, velvetleaf, and yellow nutsedge. The most sensitive weeds, including coffee senna, smallflower morningglory, and velvetleaf, were severely injured by the lowest rate when sulfentrazone was applied with surfactants. Sulfentrazone provided the highest control of yellow nutsedge with X-77. Soybeans were not severely injured by sulfentrazone applied alone, but 55% foliar injury occurred when the herbicide was applied with X-77. However, the seedlings were not killed. Sicklepod was the most tolerant of the weeds tested. In the absence of surfactants, the order of radiolabeled sulfentrazone absorption by the foliage was Centennial (5.8%) = Hutcheson (8.5%) = coffee senna (10.4%) < yellow nutsedge (17.0%) < velvetleaf (22.3%) = smallflower morningglory (24%). Sicklepod leaves did not retain droplets containing sulfentrazone when no surfactant was used. Species with the highest foliar absorption also showed the greatest phytotoxic response to the herbicide. Addition of surfactants to the spray mixture enhanced the foliar absorption and overall phytotoxicity of sulfentrazone in the weeds. An inverse relationship was detected between the foliar absorption of sulfentrazone without surfactants and the amount of cuticular wax present on the leaves. No such correlation was observed when surfactants were used. Thus, surfactants overcame the barrier to absorption imposed by the cuticular wax and, under these conditions, selectivity apparently became dependent upon species-specific cellular tolerance to sulfentrazone.

Natural products represent a vast repository of materials and compounds with evolved biological activity, including phytotoxicity. Some of these compounds can be used directly or as templates for herbicides. The molecular target sites of these compounds are often unique. Strategies for the discovery of these materials and compounds are outlined. Numerous examples of individual phytotoxins and crude preparations with weed management potential are provided. An example of research to find a natural product solution of a unique pest management problem (blue-green algae in aquaculture) is described. Finally, the problems associated with natural products for pest control are discussed.

Herbicides inhibit biochemical and physiological processes or both with lethal consequences. The target sites of these small molecules are usually enzymes involved in primary metabolic pathways or proteins carrying out essential physiological functions. Herbicides tend to be highly specific for their respective target sites and have served as tools to study these physiological and biochemical processes in plants (Dayan et al. 2010b).

Cogongrass is a troublesome, invasive weedy species with reported allelopathic properties. The phytotoxicity of different constituents isolated from roots and aerial parts of this species was evaluated on garden lettuce and creeping bentgrass. No significant phytotoxic activity was detected in the methylene chloride, methanol, or water extracts when tested at 1.0 mg ml−1. However, the total essential oil extract of cogongrass aerial parts was active. Bioactivity-guided fractionation of this extract using silica gel column chromatography led to the identification of megastigmatrienone, 4-(2-butenylidene)-3,5,5-trimethyl-2-cyclohexen-1-one (also called tabanone), as a mixture of four stereoisomers responsible for most of the activity. Tabanone inhibited growth of frond area of lesser duckweed, root growth of garden onion, and fresh weight gain of garden lettuce with 50% inhibition values of 0.094, 3.6, and 6.5 mM, respectively. The target site of tabanone is not known, but its mode of action results in rapid loss of membrane integrity and subsequent reduction in the rate of photosynthetic electron flow.