New Formulation of Clethodim/Adjuvant at Control of Grass Weeds for Soybean Crops

The use of ACCase-inhibiting herbicides without the correct addition of an adjuvant is a major cause of inefficient poaceous weed control. As such, this study aimed to assess the efficiency of a new clethodim/adjuvant formulated mixture in postemergence weed control for soybean crops. Two field experiments were conducted in the 2015/16 and 2016/17 growing seasons. A randomized block design, consisting of ten treatments with four repetitions, was used. The treatments and doses were: clethodim (108 g a.i. ha) + Lanzar® (0.5%), clethodim (108 g a.i. ha) + Nimbus® (0.5%), clethodim/adjuvant formulation at doses of (84 g a.i. ha), (96 g a.i. ha), (108 g a.i. ha), (120 g a.i. ha), (132 g a.i. ha), and (144 g a.i. ha), and a control with and without weeding. The formulated clethodim/adjuvant mixture showed high control at 7 days after application (DAA) in the 2015/16 growing season. At 28 DAA, formulation doses of 108 g a.i. ha-1 and higher exhibited superior weed control and the highest crop yields. Therefore, the use of correct adjuvant or formulated mixture is essential to increase the efficiency of clethodim herbicide.

existing mechanisms of action (Loureiro et al., 2017). These plants compete aggressively with soybean crops and can cause seed yield losses of more than 50% (Vidal & Trezzi, 2006;Barroso et al., 2010). Management practices that can lead to the selection of resistant weed biotypes include weed control using herbicides and repeatedly applying the same mechanism of action over several growing seasons (Jugulam & Gill, 2018).
Using mechanisms of action other than EPSP inhibition (glyphosate) is important to reduce the selection pressure of this herbicide on weeds. Among current herbicides, ACCase inhibitors are the best alternative to control poaceous weeds (Melo et al, 2012), which exhibit highly competitive potential with soybean crops (Barroso et al., 2010).
The occurrence of herbicide-resistant weeds in sufficient density to limit crop yields signals the need to change the management practice used (Mwendwa et al., 2018) and improving the effectiveness of herbicides. One way to enhance the performance of herbicides is the addition of corrected adjuvants on tank-mixture (Prado et al, 2016;Alves et al, 2019), although this are not active ingredients, they influence application efficiency (Maciel et al., 2014). The mineral oil, for instance, is known to increase the efficiency of ACCase-inhibiting herbicides (Costa et al., 2013), in this way, those herbicides should be always used in association with an adjuvant (Sharma & Renjith, 2016). In a study that used clethodim to control Urochloa plantaginea after soybean emergence, adding oil to the spray solution increased control from 75 to 97% (Purí ssimo, 2002). Similarly, in comparison with other adjuvants, the oil mineral presented the best efficiency in the Avena sativa control with haloxifop herbicide (Shá nchez, et al, 2018).
Achieving maximum agronomic efficiency in the postemergent management of weed species under no-till systems requires not only studies on the best application times, but combining available herbicides with adjuvants (Travlos, Cheimona & Bilalis, 2017;Prado et al, 2016)). Given that adjuvants enhance the effects of herbicides, the package inserts for all ACCase inhibitors recommend the addition of adjuvants. However, in some cases herbicides are sprayed without an adjuvant or, more frequently, in conjunction with a non-recommended adjuvant. According to the usage, the adjuvants can be divided in formulation adjuvants, when they are part of the formulation, or spray adjuvants, when they are added in the tank mix (Coalova, et al, 2014).
The apparent improvement in efficacy through the addition of adjuvants is linked to an increase in the rate of herbicide absorption (Akhter et al., 2017;Santo et al., 2017). There is not any research about the efficiency of the clethodim/adjuvant formulated mixture in weed control. As such, the aim of this study was to assess the efficiency of a new clethodim/adjuvant formulated mixture in poaceous weed control after soybean emergence and its impact on seed yield.

Experiment Site
Two field experiments were carried out in the experimental area on the Sertã o Campus of the Federal Institute of Education, Science and Technology of Rio Grande do Sul (28°02'33"S, 52°16'03"W), 705 meters above sea level, in the 2015/16 and 2016/17 growing seasons, using Distroferric Red Nitisol, whose chemical characteristics are reported in Table 1. The characteristics of the water that was used to spray is: pH 6.6, temperature 22.5°C and turbidity 2.3 NTU. The regional climate is humid subtropical (Cfa), according to the Köppen classification system.

Treatments and Experimental Design
A randomized block design, consisting of ten treatments (Table 2) with four repetitions in both growing seasons, was used. The experimental units were 2 m wide (four rows) and 4 m long. The herbicides were applied after crop and weed emergence, when plants exhibited 3 to 4 tillers. Spraying was performed using a CO2 pressurized sprayer equipped with a boom containing four ceramic spray nozzles (Micron 11002/Air) spaced 0.50 m apart and capable of administering 150 L ha -1 of spray solution.

Experiment Details
Both experiments were conducted in a no-till system with a wheat-soybean cropping sequence. Desiccation was not performed and the few dicotyledonous weeds that emerged in the experimental areas were removed by hand. Planting occurred on November 22, 2015 and December 11, 2016 using the Nidera 5959 IPRO cultivar, at a density of 350,000 seeds ha -1 and 45 cm between rows. The adjuvants were added in line with recommendations on the package insert, at 0.5% of the spray solution volume. The predominant weeds were identified as Alexandergrass (Urochloa plantaginea), Jamaican crabgrass (Digitaria horizontalis), Goosegrass (Eleusine indica) and common wheat (Triticum aestivum).

Data Collect
Weed control was assessed visually at 7, 14, 21 and 28 days after application (DAA) of the treatments, using a percentage scale from 0 to 100%, where 0 indicates no symptoms and 100% the death of all plants. Weed dry weight was evaluated at 28 DAA by collecting weed shoots from a 0.25 m 2 area per plot, which were then dried in an oven at 62º C. Seed yield was calculated based on the weight of the manually harvested seeds (two central rows of the plots, disregarding 0.5 m at each end) and extrapolated to hectares, with moisture content subsequently standardized to 13%. Climate data during the crop development stages were obtained from the Brazilian Agricultural Research Company (EMBRAPA).

Statistical Analysis
The variables were submitted to ANOVA using ASSISTAT software (Silva and Azevedo, 2002). The data were submitted to root square transformation plus one. The year was considered a random variable and the main effects of the herbicide treatment were tested for year-associated error based on treatment interaction. The data were grouped over the years when significant year-x-treatment interaction did not occur. The difference between the means was determined by Tukey's test at 5% probability and graphs were generated using Sigmaplot software (version 11.0).

Climatic Condition
The climate in both growing seasons was characterized as El Niño, with above average rainfall for southern Brazil. In the 2015/16 season, January was marked by a period of drought with irregular rainfall at levels below historic records, whereas rainfall in 2016/17 ISSN 2166-0379 2020 was normal and well-distributed over 10-day periods during soybean development (Figure 1).  ISSN 2166-0379 2020 growing season; S15/16 = treatment spraying 2015/16 growing season; S16/17 = treatment spraying 2016/17 growing season.

Weed Control
Analysis of variance for toxicity caused by the herbicides at 7, 14, 21 and 28 DAA showed interaction between the herbicides and growing seasons. In the 2015/16 crop, the treatments with clethodim/adjuvant showed greater toxicity to weeds at 7 and 14 DAA, indicating faster control. Herbicide-related symptoms were slower to emerge in the 2016/17 season compared to 2015/16, with significant control similar to the first year of the experiment only observed at 14 DAA for the highest formulated mixture doses, and at 21 and 28 DAA for all the herbicides (Figure 2), this difference is likely climate related. The effects of the herbicides intensified in 2015/16 due to a drought, with periods of higher insolation and temperatures, these climatic conditions heighten the efficacy of ACCase inhibitors (Cieslik, et al, 2013). The highest level of insolation improving metabolic activity, especially photosynthesis, which influences by increasing the translocation of the product within the plant; regarding to temperature, when the plants are exposed to lower temperature increase the leaf wax and decreased metabolism, resulting in lower absorption and translocation (Cieslik et al, 2013). The use of ACCase inhibitors under suboptimal conditions and doses has prompted weed mutations, making them resistant to these herbicides (Saini et al., 2015).
A comparison between the herbicides at 7 DAA in the 2015/16 growing season indicated control greater than 60% for all treatments except clethodim+Nimbus®, this can be explained for the fact that each adjuvant act in different ways (Cunha et al, 2017). In the growing season 2016/17 no differences between herbicides at 7 DAA. At 14 and 21 DAA, clethodim/adjuvant doses greater than 96 g a.i. ha -1 showed superior control compared to the remaining treatments, and greater efficiency in the 2016/17 season ( Figure 2). However, although weed growth stops immediately after ACCase inhibitor application, symptoms of the herbicide are not immediately apparent because of its slow translocation and action in the meristems (Maciel et al., 2014).
With respect to toxicity, at 28 DAA doses larger than 108 g a.i. ha -1 of the clethodim/adjuvant formulated mixture exhibited control greater than 90%, a significant difference in relation to the other treatments ( Figure 2). In Lolium multiflorum control, symptom evolution is related to the herbicide (clethodim) dose applied, that is, the higher the dose, the more rapidly symptoms progress (Vargas et al., 2006). There were no differences between herbicides in the 2016/17 crop (Figure 2). ISSN 2166-0379 2020  Means followed by the same lower case letter do not differ within the same growing season according to Tukey's test at 5%. Means followed by the same upper case letter do not differ in different years according to Tukey's test at 5%.

Number of Weeds and Dry Weight
Differences were observed between the herbicides for the number of weeds variable. The ISSN 2166-0379 2020 factor years and herbicide x year interaction showed no significant differences. The same was true for the variables weed dry weight and yield, therefore, the data on these variables are presented as an average for the two-year growing seasons. The smallest number of weeds was recorded for the clethodim/adjuvant treatment at doses of 120, 132 and 144 g a.i. ha -1 , indicating better control than the herbicides that require an adjuvant in the spray solution ( Figure 3). This demonstrates the importance of using the correct adjuvant with ACCase inhibitors, as is the case of the clethodim/adjuvant formulated mixture, which shows the correct combination of adjuvant type and dose. The formulated mixture also avoids mistakes on the tank mix, since there is not enough information about this (Gazzieiro, 2015), the farmers usually do not have the preparing to perform this technique. The clethodim/adjuvant herbicides at 120, 132 e 144 g a.i. ha -1 exhibited fewer weeds at the end of the experiment in relation to the other treatments (Figure 3). These results indicate the importance of the adjuvant in improving the efficiency of ACCase inhibitors and, particularly, that adjuvants differ. The importance and efficiency of adjuvants is especially evident when they are used for weed control under unfavorable conditions (Maciel et al., 2014). The use of oils as adjuvants in spray solutions with systemic postemergent herbicides can increase weed absorption of the active ingredient, improving control (Mendonç a, 2003). Means followed by the same lower case letter do not differ within the same assessment period according to Tukey's test at 5%.

Journal of Agricultural Studies
Analysis of weed dry weight indicated low values in treatments with small final weed numbers, with the lowest dry weight recorded for the clethodim/adjuvant herbicide at doses ISSN 2166-0379 2020 over 96 g a.i. ha -1 (Figure 4). Analysis of clethodim toxicity in the control of Cynodon dactylon revealed symptoms for all the formulations/concentrations used (120 and 240 g L -1 ). However, there was a greater decline in weed dry weight for the 120 g L -1 formulation at 140 g a.i ha-1 than the 240 g L -1 formulation at 140 g a.i. ha -1 (Nandula et al., 2007). Means followed by the same lower case letter do not differ within the same assessment period according to Tukey's test at 5%.

Soybean Yield
The yield results demonstrated that poaceous weed infestation of the soybean crop prompted a 2064 kg ha -1 decline in seeds compared to the weed-free control ( Figure 5). The weed interference in soybean during all the crop cycle reduces yield 73 to 94% in different sowing times (Zandona, et al, 2018). Considering a 5% tolerable yield decline, the soybean crop was negatively affected by weed infestation starting at 25 DAE (PAI) (Benedetti et al., 2009). The inefficient poaceous weed control provided by the control without herbicide and resulting increase in weed interference with the soybean crop caused a significant decrease in seed yield (600 to 1400 kg ha -1 ) (Purí ssimo, 2002).
The clethodim+Nimbus® treatment was the only herbicide whose yield differed from that of the weed-free control ( Figure 5). Emphasizing one more time the importance of using the correct adjuvant for each herbicide. Doses over 96 g a.i. ha -1 of the clethodim/adjuvant formulated mixture exhibited good control and an increase of 1749.61 kg ha -1 in seed yield in relation to the weed-free control ( Figure 5). Competition of weeds with crops can generate irreversible losses, with no recovery of development or yield, even after the elimination of it (Agostinetto et al, 2014). Moreover, increase in the period when the crop remains with weed interference, greater the damage (Zandona et al, 2018).
This gain is related to the faster action of the clethodim/adjuvant herbicide in relation to the other formulations, thereby reducing weed interference in this economically important crop. The absorption and translocation of the clethodim herbicide with carbon 14 (14C) showed that absorption of the 120 g L -1 formulation 72 hours after application was 70% higher than that of the 240 g L -1 formulation (33%). Thus, the lethal dose of clethodim was more frequently translocated in plants treated with the 120 g L -1 formulation compared to those that received the 240 g L -1 formulation (Nandula et al., 2007).
Figure 5. Soybean seed yield (ha -1 ) as a function of herbicide treatments at the end of the experiment Means followed by the same lower case letter do not differ within the same assessment period according to Tukey's test at 5%.

Conclusion
The formulated mixture was efficient at controlling poaceous weeds, demonstrating superior control compared to the other clethodim formulations whose package inserts recommend the addition of adjuvants. Clethodim/adjuvant doses over 108 g a.i. ha -1 showed greater control and a smaller final number of weeds with lower dry weight values. There was an increase in soybean yield at clethodim/adjuvant dose of 96 g a.i. ha -1 or higher indicating that the formulated mixture acts more rapidly on weeds, reducing their interference with soybean crops.
In face of what was found, stands out the importance of use the right adjuvant with clethodim. ISSN 2166-0379 2020 Moreover, recommend to use the formulated mixture, clethodim/adjuvant, since it showed superior efficacy when compared to the tank mix adjuvant, besides that the formulated mixture has more security for avoiding possible mistakes, that can compromise the herbicide efficiency, when the adjuvant is add in the spray tank along with the herbicide.

Journal of Agricultural Studies
Further research is suggested using the formulated mixture in association with other herbicides to determine the effect of these mixtures.