MATERIALS AND METHODS

The study was conducted from January to April 2018 in a greenhouse, which consisted of two experiments. Plants from an area with a record of glyphosate resistance in the city of Santa Maria, Rio Grande do Sul, Brazil, were used in both experiments. Experimental units consisted of 5 L plastic pots filled with Gray-Brown Argisol. Each experimental unit contained a single Conyza spp. plant collected in the field at a standard height of 20 cm and immediately was transplanted. Plants were irrigated daily until the average height of 25 cm, when treatments were established in the different experiments.

Experiment I aimed to evaluate the efficacy of the saflufenacil herbicide in Conyza spp. plants with different injuries, and a experimental design was carried out in completely randomized design, with four replicates. Treatments were arranged in a 2x10 factorial scheme, in which factor A represented leaf integrity and factor B consisted of different levels of injury and saflufenacil application, according to Table 1. For factor B, cutting process was made using a scissors, and the different periods were used to obtain plants without regrowth (cutting at 1 Day before application (DBA)) and with regrowth (cutting at 8 DBA), simulating situations of loss of Conyza spp. shoots during harvest.

Table 1: Treatments related to Experiment I arranged in a factorial scheme.

The herbicide saflufenacil was applied at a rate of 35 g (active ingredient) a.i ha^-1, with addition of 0.5% mineral oil, at the time indicated in the Table 1. Treatments were applied with a CO₂ pressurized backpack sprayer equipped with 110.015 nozzles and calibrated to provide an application volume of 150 L ha^-1. The variables analyzed were controlled at 7, 14, and 21 days after herbicide treatment (DAT), in a percentage scale where zero (0) represents the absence of injuries, and one hundred (100) represents complete plant death. At 21 DAT, shoots were collected and placed in a greenhouse with forced air circulation at 65 °C for three days to determine the dry matter (DM) of fleabane plants.

Experiment II aimed to evaluate the translocation capacity of the herbicide saflufenacil when applied in different places in fleabane plants.It was conducted in a completely randomized design, with three replicates. Treatments consisted of the localized application of 35 g a.i. ha^-1 of the herbicide saflufenacil, with an addition of 0.5% mineral oil, in the following plant structures: leaf located close to the apex (new leaf); leaf located close to the base (old leaf); stem located close to the apex (new stem); stem located close to the base (old stem); and the region of the cut in 10 cm plants. The herbicide was deposited on plant structures using a flexible brush in a region of approximately 1.5 cm at the end of the day (7:00 PM). The variable analyzed was the percentage of necrotic area in fleabane plants at 1, 3, 5 and 7 DAT.

To measure the percentages of necrotic area, fleabane plants were photographed in all experimental units at each evaluation period. The software ImageJ® was used for image processing, where green plant parts were selected. Subsequently, non green parts were suppressed from the image, which was considered to be necrotic areas due to the action of the herbicide (Figure 1). Afterward, the percentage of the necrotic area was calculated by the difference between the total plant area and the green area, that means, without necrosis symptoms from herbicide activity, according to Ramos et al. (2015).

Figure 1: Identification and quantification of the percentage of the necrotic area of fleabane plants (Conyza spp.) after localized application of saflufenacil, indicating the original image without manipulation (A) and the selection of damage areas underlined in red color (B).

The collected data were subjected to analysis of variance by the F test (P<0.05). When statistically significant, the means were compared by the Tukey test (P<0.05) using R software (R Core Team, 2020).

RESULTS AND DISCUSSION

Analysis of variance showed statistical significance for all variables analyzed in both experiments. In experiment I, fleabane control at 7 DAT was higher than 75% in all treatments with saflufenacil application, differing from the other treatments without applying of the herbicide (Table 2). Saflufenacil proved to be an efficient alternative in the control of glyphosate-resistant fleabane biotypes and acetolactate synthase (ALS)-inhibiting herbicides, and can be mixed with herbicides with other mechanisms of action (Dalazen et al., 2015; Davis et al., 2010).

Table 2: Conyza spp. control (%) in different tissue conditions at 7, 14, and 21 days after application (DAT) of saflufenacil.

Comparing the leaf integrity of plants, the treatments in which plants were previously defoliated showed a higher control percentage at 7 DAT, except for the treatment with cutting+herbicide of 10 cm plants at 1 DBA (Table 2). Treatments with and without leaves, the most significant weed controls (higher than 90%) were observed when saflufenacil was added to treatments on defoliated plants. Moreover, approximately 20% higher control was achieved by applying the herbicide to whole plants (uncut) without leaves compared to treatment with leafy plants. This result is related to the deposition of saflufenacil on the stem of fleabane plants due to the absence of leaves. Thus, once absorbed in the stem, saflufenacil is translocated to the other parts of the plant, acting on the chlorophyll synthesis pathway (Oliveira Júnior, 2011). The green stem of fleabane plants stands out regarding the presence of this photosynthetic pigment, which can favor the mode of action of the herbicide.

The results showed that saflufenacil application on cut-leaf plants, simulating the harvest of winter cereals, was higher to the treatment of uncut-leafy plants at 7 DBA (Table 2). However, defoliated cut plants showed lower controls when compared with treatments mixed with herbicides. In this sense, fleabane plants have a high regrowth capacity after management practices with potential for recovery (Oliveira Neto et al., 2013).This regrowth capacity highlights the need to adopt alternatives that contribute to chemical and mechanical control through other practices such as crop management (Oliveira Neto et al., 2010).

At 7 DAT, the treatment with the cutting of 10 cm plants at 1 DBA plus herbicide application did not differ between plants with and without leaves, showing satisfactory control (93%). This shows greater sensitivity of 10 cm fleabane plants for saflufenacil application (Table 2). Also, 20 cm plants that were cut and received saflufenacil did not differ from whole plants that received the herbicide, regardless of leaf integrity (Table 2). Growth and development characteristics must be observed for proper control of Conyza spp. through saflufenacil. Higher plants in an advanced growth stage have a high capacity for lateral shoot emission to recover from herbicide exposure (Moreira et al., 2010). However, this study showed reasonable control of Conyza spp. plants up to 25 cm height.

After 14 and 21 DAT, the application saflufenacil in Conyza spp. plants of 10 and 20 cm was efficient in all treatments, with control equal to or greater than 98% (Table 2). Treatments without saflufenacil application generally showed a decrease in fleabane mechanical injuries from 14 DAT, differing from herbicide treatments and indicating plant recovery (Table 2). When associating saflufenacil and glyphosate, a similar study showed satisfactory control (97%) at 14 DAT of Conyza bonariensis plants with regrowth and glyphosate resistance in an apple orchard (Pereira et al., 2016). Furthermore, mechanical weed control of Eryngium horridum with 10 to 15 cm mowing in natural pasture removed the shoots of weeds; however, it did not reach lateral buds, allowing regrowth at 20 DAT (Pellegrini et al., 2007), a similar situation was observed in the present study.

Treatments with saflufenacil application decreased DM in comparison to those that have not applied herbicide in whole plants with or without leaves (Table 3). The DM of whole leafy plants decreased by 64% after saflufenacil application compared to an entire leafy plant without herbicide treatment. Cesco et al., (2019) evaluated Conyza spp. control at different stages of plant development, the saflufenacil rate of 60 g a.i. ha^-1 decreased DM by approximately 77 and 54% in fleabane plants up to 5 and 20 cm, respectively, and led to a low regrowth percentage in plants up to 20 cm height.

Table 3: Dry matter (DM) of Conyza spp. in different tissue conditions at 21 days after application (DAT) of saflufenacil.

In treatments with saflufenacil application, DM differed only between whole plants with and without leaves, with lower DM in defoliated plants (Table 3). Besides, defoliation of Conyza spp. plants decreased DM approximately by 50 to 70%, in the control treatment (whole leafy plant) and in treatments in which the plants had regrowth (cutting at 8 DBA, without herbicide application), showing statistical difference (Table 3). The results indicated the efficiency of saflufenacil in controlling Conyza spp. plants with injured leaves, for example, after harvesting damage. Moreover, the control and reduction of DM in defoliated Conyza spp. plants may be related to herbicide absorption in the plant stem.

Treatments with saflufenacil application on leaves (new and old ones) showed a necrotic area less than 20% in all evaluations (Table 4). Foliar applications of saflufenacil tend to concentrate in the meristematic regions of the plant and not translocate towards the roots (Budd et al., 2017). Furthermore, it is known that this herbicide has a weak acid character, with greater translocation via xylem and limited translocation via phloem (Grossmann et al., 2011), which may justify the low mobility from leaves. A study on saflufenacil absorption and translocation in wheat (Triticum aestivum L.) found that the absorbed herbicide showed low translocation from treated leaves, corroborating the data observed in the present study (Frihauf et al., 2010).

Table 4: Necrotic area (%) of Conyza spp. evaluated at different times after localized application of saflufenacil.

The lowest average necrotic areas were observed in the treatment where the herbicide was applied only in the region of the cutting, being lower to the other treatments at 3, 5, and 7 DAT, with a necrotic area less than 2% (Table 4). On the contrary, picloram application in the cut stem of a woody species (Tectona grandis) showed translocation via phloem, being efficient in controlling future shoots (Caldeira and Castro, 2012), which shows a difference in systemic herbicides about those with less mobility and support the importance of knowing the behavior of the applied herbicide and the target weed. In the present study, the necrotic region resulting from stem application must be related to chlorophyll in the fleabane stem, which has a photosynthetically active area. The interaction of protoporphyrin IX with oxygen in presence of light generates singlet oxygen, resulting in lipid peroxidation of membranes and interruption of chlorophyll and heme synthesis (Owen et al., 2011). Moreover, products tend to produce hydrogen peroxide and lead to rapid necrosis and wilting, disrupting cell membranes and reducing herbicide translocation (Eubank et al., 2013).

Saflufenacil application on the stem located close to the base (old stem) showed a larger necrotic area at 3, 5, and 7 DAT than the other treatments, demonstrating that this region has greater herbicide translocation capacity (Table 4). The biochemical characteristics of saflufenacil, such as pKa 4.4, octanol-water partition coefficient (Kow 2.6), and ion trapping mechanism, provide it with metabolic stability and assist in its translocation in the plant (Kleier et al., 1996). When this herbicide is applied to the basal region of Conyza spp. plants, these characteristics allow transport via xylem for a longer period before total destruction of plant structures occurs, since damage to vascular tissues occurs more slowly after absorption. Moreover, it is known that photoassimilate distribution occurs from the producing region (source) to the metabolic and/or storage regions (sink) (Taiz and Zeiger, 2017). These factors may explain the larger necrotic area when saflufenacil was applied in the region close to the base, considered a source region.

CONCLUSION

Saflufenacil is efficient in the control of Conyza spp. having a higher action in leafless and cutting plants. The saflufenacil absorption is carried out by the stem in Conyza spp. plants. The leaves showed low absorption and translocation from the located application of saflufenacil and this herbicide translocated from the stem to the leaves.