Status of Amaranthus Species in Ohio Crop Production

Thesis

Presented in partial fulfillment of the requirements for the degree Master of Science in the Graduate School of The Ohio State University

By

Samantha N. Konkle, B.S.

Horticulture and Crop Science Graduate Program

The Ohio State University

2015

Thesis Committee:

Dr. Mark M. Loux, Advisor

Dr. Laura Lindsey

Dr. Kristen Mercer

Copyright by

Samantha N. Konkle

2015

Abstract

Studies were conducted in 2012, 2013, and 2014 with the objectives of:

1) Determining the frequency and distribution of horseweed (Conyza canadensis), giant ragweed (Ambrosia trifida), and three Amaranthus species, Palmer amaranth

(Amaranthus palmeri), waterhemp (Amaranthus tuberculatus), and redroot pigweed

(Amaranthus retroflexus), in Ohio soybean fields at the end of the growing season;

2) Determining the response of Ohio Amaranthus species to herbicide site of action groups 2, 9, and 14; and

3) Developing herbicide-based management strategies for herbicide-resistant redroot pigweed.

A survey was conducted in 2012, 2013, and 2014 in 52 Ohio counties just prior to soybean harvest, and each soybean field encountered was assessed for infestation level of horseweed, giant ragweed, and Amaranthus species. Amaranthus species were the least frequent in comparison with horseweed and giant ragweed in all three years of the survey. Palmer amaranth was not observed on the survey in any of the three years, but seed samples were collected from growers and provided by agribusinesses. Waterhemp infestations were observed in four and ten fields in 2013 and 2014, respectively. Redroot pigweed infestations occurred in 34, 19, and 2 fields in 2012, 2013, and 2014,

ii respectively. There was no pattern for the distribution of these infestations throughout the state.

A greenhouse herbicide screen was conducted using Amaranthus species samples from the 2013 and 2014 surveys to determine response of populations to glyphosate (site

9), fomesafen (site 14), and imazethapyr (site 2). The experiment was conducted twice in 2013 and once in 2014. Results varied across the two 2013 experiments, as herbicides were more active in the second experiment than in the first. Redroot pigweed populations exhibited resistance to site 2 and 14 herbicides in all three experiments.

Glyphosate resistance was inconsistent between the 2013 experiments, but was observed in 2014. Redroot populations also exhibited multiple resistance for sites 2 and 14 in 2013 and 2014, and three-way resistance occurred in the second 2013 and the 2014 experiment. Waterhemp populations were resistant to sites 2 and 9 in all experiments, but resistance to site 14 was less consistent. Multiple resistance also occurred in waterhemp populations. In 2013, this was seen only for sites 2 and 9, but in 2014, multiple resistance was seen for all combinations of site 2, 9, and 14. None of the three

2013 Palmer amaranth populations were glyphosate-resistant, but several 2014 populations were. All Palmer amaranth populations in all experiments showed resistance to site 2, and resistance to site 14 was inconsistent in 2013 but was also observed in the

2014 experiment. Three-way resistance and resistance to site 2 and 14 and site 9 and 14 were observed in the 2014 populations.

Two field studies were conducted in 2013 and 2014 to assess control of redroot pigweed using a number of herbicides with residual activity, and compare control between currently available herbicide programs and prospective herbicide programs for

iii new herbicide resistance traits. The results of the residual study showed the most effective late-season control with three-way herbicide mixes, while the least effective control was provided by 2,4-D and metribuzin applied individually or together.

Preemergence treatments in the herbicide programs study mostly provided over 90% control at 21 DAT, but many treatments showed less control starting at 42 DAT. Several preemergence + EPOST and preemergence + LPOST treatments, including a preemergence application of pyroxasulfone + flumioxazin followed by various tank mixtures postemergence, provided 90% or greater late-season control. An advantage for either current herbicide programs or prospective herbicide programs was not observed in either study.

iv

Vita

January 12, 1991 ...... Born – Columbus, Ohio

December 2012 ...... B.S. Agriculture, The Ohio

State University

January 2013 to present ...... Graduate Research Associate,

Department of Horticulture

and Crop Science, The Ohio

State University

Field of Study

Major Field: Horticulture and Crop Science

Specialization: Weed Science

v

Table of Contents

Abstract ...... ii

Vita ...... v

List of Figures ...... viii

List of Tables ...... ix

Chapter 1: Literature Review ...... 1

Chapter 2: Distribution characteristics of Ohio Amaranthus spp.

Introduction ...... 16

Objectives ...... 17

Materials and Methods ...... 17

Results and Discussion ...... 19

Bibliography ...... 22

Chapter 3: Assessment of herbicide resistance in Ohio Amaranthus spp.

Introduction ...... 31

Objectives ...... 33

Materials and Methods ...... 33

Results and Discussion ...... 35

Bibliography ...... 42

vi Chapter 4: Control of Redroot Pigweed Utilizing Current and Prospective Herbicide

Technologies

Introduction ...... 49

Objectives ...... 51

Materials and Methods ...... 51

Results and Discussion ...... 53

Bibliography ...... 57

Thesis Bibliography ...... 70

vii

List of Figures

Figure 2.1 All counties surveyed in 2012, 2013, and 2014 ...... 23

Figure 2.2 Distribution of known Palmer amaranth and waterhemp populations in Ohio in

2012 ...... 28

Figure 2.3 Distribution of known Palmer amaranth and waterhemp populations in Ohio in

2013 ...... 29

Figure 2.4 Distribution of known Palmer amaranth and waterhemp populations in Ohio in

2014 ...... 30

viii

List of Tables

Table 2.1 Percentage of soybean fields infested by species in end-of-season survey of 52

Ohio Counties ...... 24

Table 2.2 Number of infestations from the survey by county and species for 2012, 2013, and 2014 ...... 25

Table 2.3 Number of populations collected/submitted for determination of herbicide resistance ...... 27

Table 3.1 Response of 2013 Amaranthus spp. population to herbicides in the greenhouse

...... 43

Table 3.2 Response of 2014 Amaranthus spp. populations to herbicides in the greenhouse

...... 45

Table 3.3 Percentage of 2013 Palmer, waterhemp, and redroot populations exhibiting resistance to site 2, 9, and 14 herbicides at 1X and 4X rates and populations exhibiting multiple resistance at 1X rates ...... 47

Table 3.4 Percentage of 2014 Palmer, waterhemp, and redroot populations exhibiting resistance to site 2, 9, and 14 herbicides at 1X and 4X rates and populations exhibiting multiple resistance at 1X rates ...... 48

ix Table 4.1 Effect of soil-applied herbicides on residual control of redroot pigweed in the absence of a crop ...... 58

Table 4.2 Effect of soil-applied herbicides on population density of redroot pigweed in the absence of a crop ...... 59

Table 4.3 Effect of soil- and foliar-applied herbicides on control of redroot pigweed in the absence of a crop ...... 60

Table 4.4 Effect of soil- and foliar-applied herbicides on population density of redroot pigweed in the absence of a crop ...... 65

x

Chapter 1: Literature Review

The Amaranthaceae family, also referred to as the Amaranth family, is part of the order Caryophyllales. The plant species in the Amaranth family are vascular, flowering dicots that reproduce by seed. The Amaranth family contains approximately 23 genera and one of those is the genus Amaranthus (USDA, 2015). The genus Amaranthus consists of close to 75 species. However, the genus Amaranthus is divided into sub- genera, Acnida and Amaranthus. There is a group of 10 species that belong to the sub- genus Acnida (Steckel, 2007). Acnida species are dioecious; therefore, the male and female flowers exist on separate plants. Sub-genus, Acnida, was formerly a genus of its own before being combined with genus Amaranthus. Amaranthus species are monoecious, so the male and female flowers are on the same plant (Mosyakin &

Robertson, 1996). While floral distribution may vary across Amaranthus species, their geographical origins are similar.

A majority of Amaranthus species are native to the United States, Mexico, Central

America, and South America. In addition, concentrated areas of species have easily become more widespread as seeds are introduced to new regions. For example,

Amaranthus species concentrated in the western U.S. have spread to the eastern U.S.

(Robertson, 1981). Most of the annual Amaranthus species were pioneers in habitats such as riverbanks, shores, marshes, beaches, and canyons (Sauer, 1967). Historically,

1 amaranth seeds were gathered then toasted and milled for food, and the shoots and leaves of amaranth plants were prepared as greens (Sauer, 1967). Native American tribes used

Amaranthus species extensively as a food source (Steckel, 2007). However, the same

Amaranthus species humans used for food can be poisonous to cattle when grown in dry conditions. This is caused by a build-up of nitrates in the plant that convert to nitrites in the stomach of cattle (Burrouis & Tyrl, 2001). Grain amaranths still exist and are grown, but several Amaranthus species are problematic agronomic weeds (Sauer, 1967). Palmer amaranth (Amaranthus palmeri), common waterhemp (Amaranthus rudis)/tall waterhemp

(Amaranthus tuberculatus), and redroot pigweed (Amaranthus retroflexus) are among the common weedy Amaranthus species in the United States.

Palmer amaranth is one of the few dioecious Amaranthus species that belongs to the sub-genus Acnida (Steckel, 2007). It is native to the southwestern United States and northwestern Mexico (Sauer, 1957). Because Palmer amaranth is a dioecious plant, it is an obligate out-crosser (Steckel, 2007). Identifying characteristics of Palmer include its long petioles that are often as long as or longer than the leaf itself, hairless stem and leaf surface, and occasionally a white v-shaped variegation on the leaf. The flowering structure is a straight spike that is typically one to two feet in height (Horak et al., 1994).

Common and tall waterhemp both belong to the sub-genus Acnida, as they are among the dioecious Amaranthus species. This also makes waterhemp an obligate out- crosser (Steckel, 2007). Both species are native to the Midwest United States; however, common waterhemp was mostly found west of the Mississippi River while tall waterhemp was to the east of the Mississippi (Sauer, 1957). It has been assumed that these species are very closely related and can easily hybridized with one another making

2 it difficult to differentiate between the two species. However, in 2001, Pratt and Clark made the suggestion that there are not two separate waterhemp species, but rather a single species that differs along a morphological gradient across geographic locations (Pratt &

Clark, 2001). Mature waterhemp plants have hairless stems and leaves, similar to Palmer amaranth. However, waterhemp leaves are more slender and lanceolate-shaped compared to other Amaranthus species. The flowering structures are also more branched and slender (Horak et al., 1994).

Redroot pigweed is native to the central and eastern United States and can also be found in Mexico and Canada (Sauer, 1967). Contrary to waterhemp and Palmer amaranth, redroot pigweed is a monoecious Amaranthus species. Another differentiation factor for redroot pigweed from waterhemp and Palmer amaranth is that redroot pigweed has hair on its stem and leaves. The petioles are shorter relative to Palmer amaranth. The flowering structure is branched, but the branches are very compact (Horak et al., 1994).

Other weedy Amaranthus species include smooth pigweed (Amaranthus hybridus), Powell amaranth (Amaranthus powelli), and spiny amaranth (Amaranthus spinosis). Although there are taxonomical differences among the weedy Amaranthus species, they do possess general similarities. Amaranthus species are known for their prolific seed production (Sauer, 1967). Their seed dispersal mechanisms are also shared, as Amaranth seeds are spread via gravity, wind, water, animals, humans, and agronomic practices (Ward et al., 2013). Amaranthus species are also able to hybridize with each other, which encourages gene flow between species (Franssen et al., 2001). However, differences among physiological features still exist.

3 A study in Missouri compared the growth of several Amaranthus species where emergence times, plant heights, plant biomass, and seed numbers were measured. Palmer amaranth and redroot pigweed emerge more quickly than waterhemp. Palmer amaranth also had the greatest height at 208 cm. Palmer also had the greatest biomass among the three species. However, all three species produced over 250,000 seeds per plant (Sellers,

2003). The emergence of Amaranthus species is highly dependent on temperature.

Germination of Palmer amaranth, redroot pigweed, and waterhemp is not observed at day/night temperatures of 15/10 C. At 25/20 C, waterhemp germination was at its maximum, and at 35/30 C, redroot pigweed and Palmer amaranth germination peaked.

Plant death is observed at 45/40 C (Guo & Al-Khatib, 2003). While all three species are agriculturally important weeds, it is evident that Palmer amaranth has some competitive advantages. Sauer (1957) suggests that Palmer amaranth is one of the most successful invasive weeds in artificial habitats, which includes habitats prepared for agricultural use.

Soybeans (Glycine max) are a major U.S. oilseed crop, as they are the second most abundant crop next to corn (Zea mays). According the to USDA, over 76.4 million acres of soybeans are planted in the U.S. Improvements in genetics and management have allowed increases in soybean yields (Singh & Hymowitz, 1999). However, there are yield-limiting factors that must be managed. Among crop pests, weeds pose the biggest threat to yield loss (Oerke, 2006). The estimated annual cost of weeds in the

United States is $26.4 billion (Neve et al., 2009). Weeds not only affect the yield of crops, but also the quality (Gibson et al., 2007). Methods of weed control include chemicals, mechanical control, manual weeding, and other cultural practices (Oerke,

2006).

4 Presence of Palmer amaranth in the southern U.S. is evident, but Ohio and other northern states are now being threatened by Palmer (Heap, 2013). In 2013, 4.5 million acres of soybeans were planted in Ohio (NASS, 2013). Soybean acreage increased in

2014 to 4.85 million acres (NASS, 2014). As of 2012, at least one Palmer amaranth population was confirmed. There are a few proposed mechanisms for Palmer amaranth’s spread into the north. Cottonseed products are often used in dairy feeds, and Norsworthy et al. (2009) confirmed that some cottonseed products are contaminated with Palmer amaranth seed; therefore, it is possible that contaminated cottonseed products being fed to livestock are a source of infestation via livestock manure. Another proposed mechanism of spread involves a USDA program called Conservation Reserve

Enhancement Program (CREP). This program involves retiring agricultural land in exchange for government payments. As most of the retired areas are seeded, it is proposed that the seed used may be contaminated with Palmer amaranth seed. These mechanisms are only proposed and have not been confirmed.

Palmer amaranth has become an important agricultural weed in the southern

United States for soybean producers. In 1995, Palmer was not listed in the top five most troublesome weeds, but by 2000; it emerged as the second most troublesome weed for the south. As of 2009, Palmer remained the second most troublesome weed (Webster &

Nichols, 2012). According to Ward et al. (2013), Palmer amaranth went from being of little to no threat to being on of the widespread and issue-ridden weeds in less than 20 years. States that have been experiencing issues with Palmer amaranth in soybean fields include Arkansas, Kansas, Tennessee, North Carolina, South Carolina, Georgia,

Alabama, Missouri, Mississippi, and Virginia (Heap, 2013). Although Palmer amaranth

5 is known for causing problems in the south, some states further north have confirmed the presence of Palmer amaranth populations. Michigan, Illinois, Indiana, Delaware, and

Ohio are among those states (Heap, 2013). Much of the literature concerning Palmer amaranth is focused on the weed in soybean systems; however, it is important to note that

Palmer interferes with other crops, such as cotton, maize, and peanut (Heap, 2013).

The consequences producers are facing in the south as a result of Palmer amaranth include extreme soybean yield losses and complete losses. A study by Bensch et al. (2003) examined the effects of Palmer amaranth, redroot pigweed, and waterhemp on soybean yields. It was determined that soybean yield loss was the greatest when the

Amaranthus species were seeded at the same time as soybeans and weed density was at 8 plants per m-1 of row. Maximum yield losses when weed density was at 8 plants per m-1 at 78%, 56%, and 38% for Palmer amaranth, waterhemp, and redroot pigweed, respectively (Bensch et al., 2003). When Palmer is growing without soybeans, it accumulates a greater biomass of up to 970 grams per plant than when it is competing with soybeans; however, it is evident that even at lower biomass of 60-90 grams per plant, Palmer amaranth is able to cause severe yield losses in soybeans (Monks & Oliver,

1998). In some extreme cases, soybean fields can be a complete loss as a result of abandoned fields. In these cases, the expected yield will be so poor that the crop is not worth harvesting, or the infestation will cause difficulty for harvesting equipment to get through the field (Smith et al., 2000).

There are several factors that play a role in Palmer amaranth’s ability to be so competitive in soybean systems. Populations are able to grow very quickly as a result of advantageous biological and physiological traits, as well as developed traits.

6 Herbicide resistance is a large contributor to Palmer amaranth’s ability to compete with soybeans, therefore allowing populations to grow. Currently in the United States,

Palmer amaranth is known to be resistant to five herbicide sites of action—glycines,

ALS-inhibitors, HPPD-inhibitors, photosystem II inhibitors, and dinitroanilines.

Glyphosate resistance is that most frequently occurring herbicide resistance in Palmer amaranth (Heap, 2013). In 1996, glyphosate-resistant soybeans were released, and growers rapidly adopted the technology. By 2006, 89% of the soybeans grown in the

United States were glyphosate-resistant varieties. Therefore, producers were utilizing glyphosate to manage weed populations in 89% of soybean fields (Owens, 2008). The continued overuse of glyphosate has influenced shifts in weed populations by placing selection pressure on populations treated with glyphosate. Weeds that exhibit resistance to glyphosate thrive and reproduce in fields that are planted with glyphosate-resistant soybean varieties (Owen, 2008). Currently, Palmer amaranth is the second most important glyphosate-resistant weed next to Conyza Canadensis (Beckie, 2011).

However, some producers will continue to apply glyphosate to fields where glyphosate- resistant weeds exist. This only amplifies the issue and allows the resistant populations to keep growing.

Typically, a shift in traits, such as herbicide resistance will cause decreased fitness of a population in the absence of the particular mode of action that population is resistant to. Baucom and Mauricio (2004) found that glyphosate-resistant morning glory exhibited decreased fitness in situations where glyphosate resistance was not applied in contrast to when glyphosate was applied. However, this is not true for all species and modes of action. Sibony and Rubin (2003) found that some Amaranthus species tolerant to ALS-

7 inhibiting herbicides did not display a decrease in fitness. This can complicate trying to control resistant populations even with different modes of action. Resistance to multiple modes of action also occurs in Palmer amaranth populations (Heap, 2013).

Palmer amaranth is an obligate out-crosser and closely related to other

Amaranthus species. Therefore, Palmer amaranth has the ability to hybridize with other

Amaranthus species. This causes gene flow, and as a result, herbicide resistance mechanisms can be transferred via hybridization (Gaines et al., 2011). Although the study by Gaines et al. that observed this possibility also showed that it does not happen at a high frequency, it is still possible. This is a threat, as Amaranthus populations that exist without resistance can receive genes to confer resistance. This would mean that instead of worrying to control one herbicide-resistant Amaranthus species, there is possibility of having to control several.

Other advantageous factors playing a role in Palmer amaranth’s ability to compete with soybeans include a rapid growth rate, prolific seed production, and an extended period for emergence. Palmer amaranth performs C4 photosynthesis, and its rate of photosynthesis is very high relative to other C4 species, which allows faster growth (Ehleringer, 1983). Horak & Loughin (2000) found that Palmer produces more biomass than other Amaranthus species and has a higher growth rate and leaf area compared to other Amaranthus species. Accoring to Ward et al. (2013), the optimal time for herbicide application is before Palmer plants are 8 cm tall; however, if the window is missed, many producers are tilling up the field and replanting. Palmer’s ability to grow rapidly gives it the advantage of escaping the optimal control window and out-compete soybeans for essential resources (Steckel, 2007).

8 Like all Amaranthus species, female Palmer amaranth plants produce an abundance of seed. Plants produced between 200,000 and 600,000 seeds per plant in study done by Keeley et al. (1987). The prolific amount of seed produced contributes to the soil seed bank; therefore, contributing to population persistence. In a Palmer amaranth seed bank study, Menges (1987) determined that in non-treated conditions, the seed bank consisted of 1.1 billion seeds per ha-1. Even plots that received hand weeding and herbicide application still yielded a high volume of seed with nearly 18 million seeds per ha-1 (Menges, 1987). Palmer amaranth seeds are very small, only 1-2 cm, so they are easily dispersed (Ward et al., 2013).

Those seeds are also very viable. It has been observed that Palmer amaranth seeds retain 78% viability after six months in the soil seed bank; however, seeds became less likely to emerge with deeper burial in the soil profile (Sosnoskie et al., 2011).

Palmer amaranth seeds were more likely to emerge at depths of 2.5 cm or less (Keeley et al., 1987). Seeds buried deeper can be brought back to the surface via tillage; therefore, creating a source of re-infestation (Ward et al., 2013). The seeds are also quick to germinate and have been referred to as “opportunistic,” as they take advantage of favorable conditions quickly (Ward et al., 2013). In comparison with other Amaranthus species, Palmer amaranth had accelerated germination times. Seeds were exposed to alternating temperatures, and all of the Palmer seeds emerged within the first day while the other species took three to eight days for only 50% emergence (Ehleringer, 1987).

Seed herbivory by animals is a threat for Palmer amaranth seeds; therefore, quick emergence allows seeds to avoid that (Sosnoskie et al., 2011). In addition, Palmer amaranth seeds have an extended emergence period. Keeley et al. (1987) observed

9 Palmer amaranth emergence from March until October. The terminal date for emergence is a result of the first killing frost; therefore, it can vary (Keeley et al., 1987). This allows

Palmer to escape herbicide applications.

Currently, many of the management efforts for Palmer amaranth are focused on control in glyphosate-resistant soybeans (Ward et al., 2013). However, in a survey completed by producers, it was reported that they are not likely to reduce or eliminate their use of glyphosate in the presence of glyphosate-resistant weeds. The low cost of glyphosate for weed control in comparison to other management tactics is the influencing factor for producers (Beckie, 2011). Because producers are using glyphosate on glyphosate-resistant weeds, they typically have to implement other weed control strategies; however, this ends up costing producers more money than if they had used something other than glyphosate to begin with. Mueller et al. (2005) described proactive management as, “avoid breaking the tool, thus maintaining the tool’s effectiveness,” and reactive management as, “use the tool until it breaks, then find a new tool.” In their study done on waterhemp in Illinois, it was determined that a proactive approach to managing glyphosate-resistance with herbicides would cost growers $5 per ha-1, while a reactive approach with herbicides would cost growers $44 per ha-1 (Mueller et al., 2005). A study done on Palmer amaranth in cotton systems determined that a preventative weed control program would save growers $50 per ha-1 compared to reactive approaches (Culpepper et al., 2010).

Hand weeding and tillage are two reactive methods by which growers in the south are battling herbicide resistant weeds when herbicide applications are not effective.

According to Riar et al. (2013), the average cost of hand weeding for Palmer amaranth in

10 soybeans is between $46 and $59 per ha-1. However, some extreme cases yielded costs of up to $371 per ha-1. Typically, the cost depends on the severity of the infestation, but the choice of growers to hire labor for hand weeding depends on weed size, labor availability, and timing of weed seed production (Riar et al., 2013). Tillage increases costs for growers via the cost of fuel and time; however, herbicide resistant Palmer amaranth has decreased conservation tillage by 35% (Beckie, 2011). In addition to increasing costs, increased tillage reduces the benefits seen from utilizing conservation tillage, such as decreased erosion and compaction, increased water conservation, and improved soil quality (Price et al., 2011). Undoubtedly, Palmer amaranth management can increase costs for growers when they are not proactive. However, the yield returns on both proactive and reactive management do not differ noticeably (Beckie, 2011).

There are several proposed control methods for Palmer amaranth. Riar et al.

(2013) outlines the “best management practices” for controlling herbicide resistant weeds. Those practices include proper timing of herbicide application, planting into weed-free fields, using multiple modes of action, using full herbicide rates, preventing weed seed production, crop rotation, rotation of herbicide resistant crop traits, cover crops, narrow row spacing, altered planting date, tillage, and sanitary equipment (Riar et al., 2013). However, some of these recommendations can be difficult to follow, as they can be dependent upon weather conditions and other variables (Price et al., 2011).

Relative to chemical control of Palmer amaranth, an emphasis has been put on pre- emergent herbicides with residual activity (Price et al., 2011).

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Sibony, M. and B. Rubin. “The ecological fitness of ALS-resistant Amaranthus retroflexus and multiple-resistant Amaranthus bliotides.” Weed Research 43(1): 40-47. 2003.

Singh, R.J. and T. Hymowitz. “Soybean genetic resources and crop improvement.” Genome 42(4): 605-616. 1999.

Smith, D., R. Baker, and G. Steele. “Palmer amaranth impacts on yield, harvesting, and ginning in dryland cotton.” Weed Technology 14(1): 122-126. 2000.

Sosnoskie, L.M., A.S. Culpepper, and T.M. Webster. “Palmer amaranth seed mortality in response to burial depth and time.” Proceedings of the Beltwide Cotton Conference:1550-1552. Cordova, TN. National Cotton Council of America. 2011.

Steckel, L.E. “The dioecious Amaranthus spp.: here to stay.” Weed Technology 21: 567–570. 2007.

Steckel, L.E., T.W. Eubank, J. Weirich, B. Scott, and R. Montgomery. “Glyphosate 14 resistant Palmer amaranth control in dicamba tolerant soybean system in the midsouth.”Proceedings of the Southern Weed Science Society: 65. Las Cruces, NM. Southern Weed Science Society. 2012.

Ward, S., T.M. Webster, and L.E. Steckel . “Palmer amaranth (Amaranthus palmeri): A review.” Weed Technology 27(1): 12-27. 2013.

Webster, T.M. and R.L. Nichols. “Changes in the prevalence of weed species in the major agronomic crops of the southern United States.” Weed Science 60(2): 145- 157. 2012.

USDA, NRCS. 2015. The PLANTS Database (http://plants.usda.gov, 20 March 2015). National Plant Data Team, Greensboro, NC 27401-4901 USA.

15

Chapter 2: Distribution characteristics of Ohio Amaranthus spp.

Introduction

The Amaranth family contains several weedy members that cause issues in agronomic crop production. Palmer amaranth (Amaranthus palmeri) and waterhemp

(Amaranthus tuberculatus) are dioecious members of the Amaranth family, and redroot pigweed (Amaranthus retroflexus) is a monoecious member. Palmer is native to the southwestern United States and northwestern Mexico, and waterhemp is native to the midwestern United States. Redroot pigweed is native to the central and eastern United

States (Sauer, 1957). These three species are common agricultural weeds in U.S. soybean production.

There are several factors that contribute to the success of these weeds. Plants in the Amaranth family are known for their prolific seed production, as they can produce upwards of 250,000 seeds per plant (Sellers, 2003). Seeds also have a long germination window. Keeley et al. (1987) observed emergence of Palmer amaranth from March until

October. A rapid growth rate also aids in Amaranthus species ability to compete well with soybeans. Palmer amaranth performs C4 photosynthesis at a very high rate relative to other C4 species (Ehleringer, 1983). The presence of herbicide resistance in populations also gives pigweed species a competitive advantage. Yield losses in soybeans from Palmer amaranth, waterhemp, and redroot pigweed can be up to 78%,

56%, and 38%, respectively (Bensch et al., 2003).

16 According to a 2013 survey by the Southern Weed Science Society, Palmer amaranth is one of, if the not the most, troublesome weeds in soybean cropping systems in the southern United States (Webster, 2013). Waterhemp is an issue primarily in the

Midwestern U.S. (Hager et al., 2002). Prior to 2012, Ohio did not experience many issues with Palmer amaranth or waterhemp. Only one grower reported issues with

Palmer, and waterhemp populations in the state were isolated to only a few areas.

Redroot pigweed has been abundant in Ohio for decades, but it has also been well controlled. In 2013, 4.5 million acres of soybeans were planted in Ohio, and production increased in 2014 to 4.85 million acres (NASS, 2013; NASS, 2014). This totals a large economic impact for the state. Therefore, staying ahead of weed problems in soybeans is essential.

Surveys are useful tools to determine the distribution of weeds across large areas

(Loux & Berry, 1991). The results from this survey of Amaranthus spp. in Ohio soybean fields will provide awareness for soybean growers, as well as the ability to stay ahead of issues concerning these weeds.

Objective

Determine the frequency and distribution of horseweed, giant ragweed, and three

Amaranthus species, redroot pigweed, waterhemp, and Palmer amaranth, in Ohio soybean fields at the end of the growing season.

Materials & Methods

A survey of 52 Ohio counties was conducted in 2012, 2013, and 2014 just prior to soybean harvest to assess soybean fields for levels of infestation of Amaranthus spp., giant ragweed (Ambrosia trifida), and horseweed (Conyza canadensis). The counties

17 surveyed included any with at least 10,000 acres of soybeans, and these occurred in roughly the western two-thirds of Ohio (Figure 2.1). A total of 3994, 3644, and 3479 fields were surveyed in 2012, 2013, and 2014, respectively. Driving routes for each county were created using Google Earth. Routes were typically a diagonal stretch across each county, but large cities were avoided when possible. The same route was driven in

2012 and 2013. In 2014, new routes were created. Diagonal transects of each county were driven following the mapped routes, and every soybean field encountered was rated for infestation level. Infestation levels were based on assessments of weed populations visible above the soybean canopy from the edge of the field, and were based on the following scale: zero – weed free; one – a few single plants within a field; two – a few clusters of plants; three – many dense clusters of plants or widespread infestation. Fields rated a “two” or “three” were considered to be “infested” for the purpose of collecting

Amaranthus spp. seed samples. When infested fields were encountered, they were assigned a field number, and GPS coordinates were determined using a handheld Garmin

Oregon 450t. In 2012, seed samples were collected from fields rated a “three.” In 2013 and 2014, seed samples were collected from infested fields – those rated a “two” or

“three.” Additional samples were collected in 2014 from fields that were rated a “one.”

The seed sample for each field consisted of approximately 10 to 15 seed heads, which were combined into one mixed sample. In addition to identifying fields and collecting seed samples through the survey, growers and agribusiness were also asked, via newsletter, to provide information and/or seed samples from infested fields in their area.

Seed samples from the survey and those submitted by growers were used for a subsequent greenhouse study screening for herbicide resistance. The information

18 collected from the survey and from growers was used to generate maps displaying where various Amaranthus species are present throughout the state of Ohio.

Results & Discussion

Horseweed was overall the most abundant weed in soybean fields in early fall.

Horseweed infestations were observed in 10% of the fields averaged over the three years, while giant ragweed infestations occurred in 4.8% of the fields, and Amaranthus species occurred in less than 1%. Horseweed infestations were more frequent in 2012 and 2014 compared with giant ragweed, but were less frequent than giant ragweed in 2013.

Horseweed infestations occurred in 13%, 5.6%, and 7.1% of all soybean fields assessed on the survey in 2012, 2013, and 2014, respectively (Table 2.1). Horseweed was distributed widely throughout the state, occurring in 51, 50, and 49 of 52 counties in

2012, 2013, and 2014, respectively. The counties without horseweed infestations varied among years (Table 2.2).

In 2012 and 2014, giant ragweed infestations were seen more frequent than pigweed species, but less frequent than horseweed. In 2012, 82 giant ragweed infestations were seen in 34 counties, and in 2014, 123 giant ragweed infestations occurred in 31 counties (Table 2.2). In 2012 and 2014, giant ragweed infestations were present in 2.1% and 3.4% of all fields assessed in the survey; however, in 2013, there was an increase with giant ragweed infestations being found in 9% of all fields surveyed

(Table 2.1). Giant ragweed infestations were present in 44 counties in 2013 (Table 2.2).

Although there was an increase in giant ragweed infestations and their locations for 2013, there did not seem to be a pattern in the distribution.

19 While horseweed and giant ragweed infestations occurred throughout much of

Ohio, pigweed species infestations occurred much less frequently and were more narrowly distributed. Although redroot pigweed is known to be abundant throughout the state, only 34, 19, and 2 infestations were found at the end of the growing season in 2012,

2013, and 2014, respectively (Table 2.2). Four waterhemp infestations were found in

2013, and 10 were found in 2014. Like redroot pigweed, waterhemp infestations totaled less than 1% of all soybean fields surveyed all three years (Table 2.1). Palmer amaranth infestations were not observed on the survey in any of the three years it was conducted.

There is no distinct distribution pattern for redroot pigweed in Ohio, likely because it exists throughout the state but is typically well controlled by herbicides. In 2013, most of the waterhemp populations were isolated in the western part of the state near the Indiana border, but in 2014, the waterhemp populations were mostly in west central Ohio.

Overall, the distribution of waterhemp expanded from year to year. However, all

Amaranthus species combined occurred in fewer areas of the state relative to horseweed and giant ragweed. It is evident that season-end horseweed and giant ragweed infestations occur more frequently and are more widely distributed throughout Ohio compared to pigweed species. This is probably due more to the relative ease of controlling pigweed compared with horseweed and giant ragweed, than a lack of abundance of pigweeds in certain areas.

A total of 12, 34, and 52 Amaranthus seed samples were collected in 2012, 2013, and 2014, respectively (Table 2.3). These totals include seed samples collected from the survey and those submitted by growers. In 2012, six samples were collected on the survey, and six samples were submitted. Only one of these was waterhemp, and the rest

20 were redroot pigweed. In 2013, of the 34 total samples collected, 23 were collected on the survey – four waterhemp and 19 redroot pigweed. Eleven of the 2013 samples were submitted by growers – six Palmer amaranth, one waterhemp, and four redroot pigweed.

Of the 52 total samples from 2014, 25 were from the survey and 27 were submitted. The survey samples included 14 waterhemp, and 11 redroot pigweed. The submitted samples included 11 Palmer amaranth, eight waterhemp, and eight redroot pigweed. The increase in samples from year to year does not necessarily indicate an increase in populations throughout the state. Some of the increase can be attributed to increasing awareness of these weeds, therefore more submissions. In addition, in 2012 we collected only from fields rated as densely infested (rating of “3”), while in 2013, we collected from fields that were fields that were moderately or densely infested (rating of “2” or “3”). In 2014, we collected from any field with essentially any amount of pigweed plants. Over the three years, waterhemp and Palmer populations became more frequent and were no longer isolated to only a few areas of the state, but there was no distinct pattern for the distribution of these species (Figure 2.2, 2.3, 2.4).

21

Bibliography

Bensch, Curtis N., Michael J. Horak, and Dallas Peterson . “Interference of redroot pigweed (Amaranthus retroflexus), Palmer amaranth (A. palmeri), and common waterhemp (A. rudis) in soybean.” Weed Science 51(1): 37-43. 2003.

Ehleringer, J. “Ecophysiology of Amaranthus palmeri, a Sonoran Desert summer annual.” Oecologia 57:107–112. 1983.

Hager, Aaron G., Wax, Loyd M., Stoller, Edward W., Bollero, Germán A. (2002). Common waterhemp (Amaranthus rudis) interference in soybean, Weed Science 50(5): 607-610.

Keeley, P.E., C.H. Carter, and R.J. Thullen. “Influence of Planting Date on Growth of Palmer Amaranth (Amaranthus palmeri).” Weed Science 35(2): 199-204. 1987.

Loux, Mark M. & Berry, Mary Ann. (1991). Use of a grower survey for estimating weed problems, Weed Technology 5(2): 460-466.

National Agricultural Statistics Service (NASS). “Soybeans – Acres Planted 2013.” (http://quickstats.nass.usda.gov/results/3D229048-DC77-3206-8A83- 47E5BCD98F2A, 17 July 2015). USDA.

National Agricultural Statistics Service (NASS). “Soybeans – Acres Planted 2014.” (http://quickstats.nass.usda.gov/results/3D229048-DC77-3206-8A83- 47E5BCD98F2A, 17 July 2015). USDA.

Sauer, J.D. “Recent migration and evolution of the dioecious Amaranths.” Evolution 11(1): 11-31. 1957.

Sellers, B.A., R.J. Smeda, W.G. Johnson, J.A. Kendig, and M.R. Ellersieck. “Comparative growth of six Amaranthus species in Missouri.”Weed Science 51(3): 329-333. 2003.

Webster, Theodore M. Weed Survey – Southern States, 2013 Proceedings, Southern Weed Science Society (2013) 66: 275-287.

22

Coun%es(surveyed(

Figure 2.1 All counties surveyed in 2012, 2013, and 2014.

23

Table 2.1 Percentage of soybean fields infested by species in end-of-season survey of 52 Ohio Counties Year Total Redroot Palmer Waterhemp Giant Horseweed Fields ragweed 2012 3994 0.8% 0% 0% 2.1% 13% 2013 3644 0.6% 0% 0.1% 9% 5.6% 2014 3479 0.08% 0% 0.2% 3.4% 7.1%

24

Table 2.2 Number of infestations from the survey by county and species for 2012, 2013, and 2014 Pigweed spp. Giant ragweed Horseweed County 2012 2013 2014 2012 2013 2014 2012 2013 2014 Allen 1 4 2 1 3 7 Ashland 2 1 1 12 5 1 Auglaize 1 1 2 2 3 Brown 3 1 3 23 8 8 Butler 5 5 5 8 Champaign 2 1 6 3 10 17 8 Clark 2 1 1 4 4 6 12 13 Clermont 7 4 2 Clinton 1 17 13 12 11 Crawford 1 1 1 9 3 3 9 1 Darke 1 6 1 10 5 5 Defiance 3 1 4 7 7 6 Delaware 3 16 5 10 6 16 Erie 6 2 12 4 3 Fairfield 1 1 8 1 10 2 1 Fayette 2 2 3 4 16 6 4 Franklin 2 1 3 11 2 Fulton 3 2 3 12 16 4 Greene 2 1 2 5 1 10 10 Hancock 2 1 27 6 2 Hardin 3 4 7 2 17 10 3 Henry 1 2 23 7 4 Highland 3 2 31 5 8 Huron 1 1 1 11 4 4 Knox 3 1 6 8 3 Licking 1 1 1 6 5 1 6 2 Logan 1 3 6 6 7 11 Lorain 2 1 2 1 11 3 1 Lucas 1 9 1 Madison 1 5 22 4 16 10 7 Marion 1 1 5 13 5 9 12 8 Mercer 1 1 1 2 9 3 1 Miami 1 1 1 4 7 13 14 14 Montgomery 1 4 8 7 7 3 Morrow 1 4 7 1 5 6 3 Continued 25 Table 2.2: Continued Ottawa 1 3 3 3 7 Paulding 1 3 7 4 3 Pickaway 1 1 1 2 4 9 6 2 Preble 1 2 9 8 23 3 5 Putnam 11 5 3 Richland 4 4 1 18 2 5 Ross 2 1 6 10 7 Sandusky 2 6 1 4 15 8 7 Seneca 1 4 25 5 2 Shelby 4 3 4 1 4 13 Union 1 2 1 4 4 9 2 4 Van Wert 4 6 3 1 Warren 1 5 5 9 Wayne 2 1 10 5 5 Williams 1 2 5 2 17 8 2 Wood 1 1 1 6 13 4 Wyandot 3 1 10 4 3 Totals 34 23 12 82 205 123 544 330 259

26

Table 2.3 Number of populations collected/submitted for determination of herbicide resistance Year Redroot Waterhemp Palmer 2012 11 1 0 2013 23 5 6 2014 19 22 11

27

2012$

Waterhemp) Palmer) Waterhemp)and)Palmer)

Figure 2.2 Distribution of known Palmer amaranth and waterhemp populations in Ohio in 2012

28

2013%

Waterhemp) Palmer) Waterhemp)and)Palmer)

Figure 2.3 Distribution of known Palmer amaranth and waterhemp populations in Ohio in 2013

29

2014%

Waterhemp) Palmer) Waterhemp)and)Palmer)

Figure 2.4 Distribution of known Palmer amaranth and waterhemp populations in Ohio in 2014

30

Chapter 3: Assessment of herbicide resistance in Ohio Amaranthus spp.

Introduction

The Weed Science Society of America (WSSA) defines herbicide resistance as,

“the inherited ability of a plant to survive and reproduce following exposure to a dose of herbicide normally lethal to the wild type. In a plant, resistance may be naturally occurring or induced by such techniques as genetic engineering or selection of variants produced by tissue culture or mutagenesis” (1998). In addition to being resistant to a single herbicide, cross-resistance and multiple resistance can also occur in plant populations. Cross-resistance is when plants are resistant to more than one herbicide from different chemical families that act at the same site of action. Multiple resistance in when a plant has resistance to herbicides with different modes of action due to having more than one resistance mechanism (Vencill et al., 2012). Herbicide mode of action refers to how a herbicide works, and herbicide site of action refers specifically to where a herbicide works within a plant.

Palmer amaranth (Amaranthus palmeri), waterhemp (Amaranthus tuberculatus), and redroot pigweed (Amaranthus retroflexus) all have confirmed cases of herbicide resistance in the United States. Palmer amaranth is reported to have resistance to site of action group 9 (glyphosate), 2 (ALS inhibitors), 27 (HPPD inhibitors), 5 (photosystem II inhibitors), and 3 (microtubule inhibitors) herbicides. Multiple resistance across up to

31 three sites of action has also been reported in Palmer amaranth. Waterhemp has also been reported to have resistance to site of action group 9, 2, 27, and 5. In addition, resistance to site of action group 14 (PPO inhibitors) has also been reported, and multiple resistance across up to four sites of action has been reported for waterhemp. Resistance to site of action group 5 and 2 has been reported in redroot pigweed. Some redroot populations have been reported as having multiple resistance to both groups 5 and 2 but this has not occurred in Ohio. There have been three confirmed reports of herbicide resistant Amaranthus species in Ohio. Waterhemp was reported to be ALS-resistant in

1996 and glyphosate-resistant in 2008. In 2010, Palmer amaranth in Ohio was reported to be glyphosate-resistant. No reports of resistance in redroot pigweed have been made

(Heap, 2015).

Herbicide resistance in weeds is typically driven by the frequent use of herbicides with the same site of action (Beckie, 2006). With growing use of herbicide-resistant crop technology, many farmers opt to repeatedly use herbicides with the same site of action because it is simple, inexpensive, and saves time; however, when herbicide resistant weeds develop, many negative consequences arise. Yield loss, crop failure, and increased production costs are all effects of herbicide resistant weeds (Vencill et al.,

2012). Although it is complicated to calculate costs of herbicide resistant weeds to farmers, prevention typically costs less than fully developed resistance (Orson, 1999).

Therefore, it is important to monitor suspect populations. Because Palmer amaranth populations in Ohio are likely migrating from the southern U.S. where resistance has been confirmed, these populations and their herbicide resistance characteristics need to be monitored.

32

Objective

Determine the response of Ohio populations of Amaranthus species – Palmer amaranth, waterhemp, and redroot pigweed – to herbicide site of action groups 2, 9, and

14.

Materials & Methods

Seed samples collected from the soybean survey and submitted by growers in

2013 were placed in a cold storage room at approximately 4° C. A total of 34 samples were collected and stored, which included 19 redroot pigweed, six waterhemp, and nine

Palmer amaranth samples (Table 3.1). Seed heads were threshed by hand, and seed was cleaned using sieves. Each sample was assigned a number for labeling purposes, and seeds were placed in plastic vials and put back into cold storage. Of the 34 total samples, three samples did not have sufficient seed supply due to rotting and immaturity. Prior to sowing seeds, plastic pots with a diameter of 12.7 cm and volume of 1.18 L were filled with Fafard 3B Mix potting media and placed on three separate greenhouse benches.

Seeds were sprinkled on the surface of the potting media in each pot, and an additional one cm of media was layered on top of the seeds. Pots were organized in an RCBD with four replications, where plant height was the rationale for grouping into replications.

Each population was randomized throughout the greenhouse on a weekly basis for the duration of the experiment. Soil moisture was maintained using an automated sprinkler system, three times a day for one minute at a time, and supplemented with watering by

33 hand as needed. Metal halide lamps were used for supplemental lighting providing a 16- hour daylength.

Herbicide treatments were applied when plants reached the height of seven to fifteen centimeters. Three herbicides were applied at two different rates, and compared with a nontreated check. The treatments consisted of foliar application of glyphosate

(site 9) at 840 and 3360 g ae/ha, fomesafen (site 14) at 350 and 1400 g ai/ha, and imazethapyr (site 2) at 70 and 280 g ai/ha. These rates represent approximately the recommended use rate (1X), and four times the recommended use rate (4X).

Appropriate adjuvants were used with each product: glyphosate – ammonium sulfate

2.5% v/v; fomesafen and imazethapyr– crop oil concentrate 1% v/v and ammonium sulfate 2.5% v/v. These specific herbicides were chosen because they are typically used and are effective in the absence of resistance for controlling Amaranthus species.

Treatments were applied using an automated spray chamber calibrated to deliver a volume of 141 L/ha at a speed of 3.5 km/hr. An even flat-nozzle Teejet 8001EVS was used for the applications. Plants were visually assessed for mortality at 21 DAT.

Populations with a survival rate of 50% or greater were considered to be resistant.

This experiment was conducted twice, but a scarcity of seed for some populations resulted in the inclusion of fewer populations in the second compared with the first experiment. Twenty-five populations were included in the second experiment – 19 redroot pigweed, four waterhemp, and two Palmer populations.

Seed samples collected on the driving survey and submitted by growers in 2014 were placed in a cold storage room at approximately 4° C. A total of 52 seed samples were collected and stored, which included 10, 22, and 20 Palmer amaranth, waterhemp,

34 and redroot pigweed populations, respectively (Table 3.2). The same methods used for the 2013 experiments were used for this experiment. The 2014 experiment was initiated in March to ensure target internal greenhouse temperatures were maintained throughout the majority of the experiment. Of the 52 total populations, 12 were excluded from the experiment due to insufficient seed supply or poor seedling emergence. Of the treated populations, seven were Palmer amaranth, 14 were waterhemp, and 19 were redroot pigweed. The 2014 experiment was not repeated due to time limitations.

Results & Discussion

For the 2013 screen, herbicides appeared to be overall more active in the second experiment than in the first, resulting in fewer populations being characterized as resistant (Table 3.1). Differences in herbicide activity are likely due to differences in external greenhouse temperatures, which affect internal temperature trends. Both experiments were conducted during winter, but external conditions were much colder during the first, which resulted in difficulty maintaining target temperatures in the greenhouse. Higher temperatures were present for the second experiment, which may have contributed to higher levels of herbicide activity. In 2014, target greenhouse temperatures were maintained due to warmer external temperatures. The overall frequency of resistance in all three species increased from 2013 to 2014.

Redroot pigweed populations showed resistance to all three sites of action, but not necessarily in every experiment (Tables 3.1 and 3.2). For the 2013 populations, glyphosate resistance was inconsistent across the two experiments, with 40% showing resistance to the 1X rate and 9% to the 4X rate in the first experiment and no populations showing resistance to either rate in the second experiment (Tables 3.3 and 3.4).

35 However, in 2014, 53% of redroot pigweed populations showed resistance to the 1X rate of glyphosate, and 11% showed resistance to the 4X rate. Resistance to site 2 and 14 herbicides in redroot pigweed was observed in all three experiments. For the 2013 redroot populations, 50 and 74% showed resistance to the 1X rate of imazethapyr in experiment one and two, respectively, and 30 and 42% also showed resistance to the 4X rate (Tables 3.3). In 2014, 74% of the redroot populations showed resistance to site 2 at the 1X rate, and 68% also showed resistance to the 4X rate (Table 3.4). In the 2013 experiments, 65% and 32% of redroot populations showed resistance to the 1X rate of fomesafen in experiment one and two, respectively (Table 3.3). Resistance to the 4X rate of fomesafen in 2013 redroot pigweed occurred only in experiment one, for 30% of the populations (Table 3.3). However, in 2014, the frequency of redroot pigweed resistance to fomesafen increased for both rates. The percentage of populations exhibiting resistance to the 1X rate of fomesafen increased to 89%, and 37% also exhibited resistance to the 4X rate (Table 3.4). Several redroot pigweed populations exhibited resistance to multiple sites of action both years, although fewer in the second experiment of 2013 where herbicides were more active. Based on that second experiment, multiple resistance only occurred for sites 2 and 14, in 32% of the populations. This was similar to the 2014 experiment, where 57% of the redroot pigweed populations had resistance to both sites 2 and 14. Multiple resistance to sites 2 and 9 did not occur in any experiment, but 37% of the 2014 populations had resistance to all three sites (Table 3.4). Three-way resistance was evident in the first 2013 experiment, but not in the second. Glyphosate- resistance in redroot pigweed has not previously been documented in the United States

(Heap, 2015). Resistance to site 2 herbicides, or ALS-resistance, has occurred for redroot

36 pigweed populations in Arkansas, Maryland, and North Dakota. Resistance to site 14 herbicides, or PPO-resistance, has not been documented in redroot pigweed in the United

States. With regard to multiple resistance, there was resistance to sites 2 and 5 in

Pennsylvania, and resistance to sites 5 and 7 in Michigan (Heap, 2015).

All 2013 waterhemp populations exhibited resistance to site 2 at the 1X rate. In

2013, 80 and 75% of waterhemp populations also showed resistance to site 2 at the 4X rate in experiment one and two, respectively (Table 3.3). In 2014, 93% of waterhemp populations were resistant to imazethapyr regardless of rate (Table 3.4). Glyphosate- resistance was observed in waterhemp in all three experiments at both rates. In the 2013 waterhemp populations, 100 and 25% showed resistance to the 1X rate of glyphosate in experiment one and two, respectively, and 20 and 25% were resistant to the 4X rate

(Table 3.3). In 2014, all waterhemp populations were glyphosate-resistant to the 1X rate, but only 43% were resistant to the 4X rate. Resistance to site 14 was less consistent, as

20% of the 2013 populations were resistant to the 1X rate in the first experiment and none in the second experiment. However, in 2014, 36% of populations displayed resistance to site 14 at the 1X rate, and 21% also showed resistance at the 4X rate (Table

3.4). Glyphosate (site 9) and ALS (site 2) resistance in waterhemp populations has been confirmed in Ohio in the past. PPO (site 14) resistance in waterhemp has not been confirmed in Ohio, but has been other places in the United States, such as in Indiana

(Heap, 2015). Some waterhemp populations also exhibited multiple resistance. In 2013, this was only observed for sites 2 and 9, but in 2014, there were populations with multiple resistance to all combinations of sites 2, 9, and 14 except 2 and 14. Multiple resistance to all three herbicides occurred for 29% of the 2014 waterhemp populations

37 (Tables 3.3 and 3.4). This has not previously occurred in Ohio, but Illinois and Missouri have confirmed populations with these resistance characteristics. Multiple resistance to sites of action 2 and 9 and 9 and 14 has not been documented in Ohio, but has been in several other states. Multiple resistance to four sites of action has currently been documented in waterhemp (Heap, 2015).

In 2013, none of the three Palmer amaranth populations exhibited any level of glyphosate resistance (Table 3.3). We suspect that Palmer amaranth populations are migrating into Ohio from the southern United States where most populations are glyphosate-resistant, so these results were unexpected. However, in 2014, 71% of the seven Palmer populations showed resistance to the 1X rate of glyphosate, and 29% showed resistance to the 4X rate (Table 3.4). In all three experiments, 100% of Palmer populations showed resistance to site of action 2 at the 1X rate, and this also occurred for the 4X rate in 2013 experiment two and the 2014 experiment (Tables 3.3 and 3.4). ALS resistance in Palmer amaranth has been documented in several other states, but not yet in

Ohio (Heap, 2015). This could be due to the low frequency of Palmer populations in

Ohio to this point. Resistance to site of action 14 in Palmer amaranth was inconsistent across the two experiments in 2013, but 57% of populations in 2014 showed PPO- resistance at the 1X rate and 14% to the 4X rate (Tables 3.3 and 3.4). There is no documentation of PPO-resistance in Palmer amaranth thus far (Heap, 2015). As with the other two Amaranth species, Palmer amaranth did exhibit some multiple resistance. In the first 2013 experiment, no multiple resistance occurred, but 50% of populations showed two-way resistance to sites 2 and 14 in experiment two. In 2014, the incidence of multiple resistance in Palmer amaranth increased, with 29% of the populations exhibiting

38 resistance to all three sites of action. In addition, 14% of the populations showed two- way resistance for sites 2 and 14 or sites 9 and 14 (Tables 3.3 and 3.4). Several occurrences of multiple resistance to sites 2 and 9 have been confirmed in the United

States, and some of these populations were also resistant to a third site of action (Heap,

2015).

Herbicide resistance exists in redroot pigweed, waterhemp, and Palmer amaranth populations in Ohio. Resistance to high rates of herbicides and the presence of multiple resistance in Amaranthus species presents an even greater threat to Ohio soybean production. From a management aspect, resistance reduces the selection of herbicides that can be used to control these populations. Of all fields surveyed and collected from in

2013 and 2014, resistant Amaranthus species totals were as follows, depending on the year and experiment: site 2 – 0.4-0.9%; site 9 – 0.2-0.8%; and site 14 – 0.1-0.7%. There was an overall increase in resistant populations from 2013 to 2014; therefore, monitoring and managing resistant or suspect populations is crucial to preventing this becoming a trend. As there is no distinct pattern for the distribution of Palmer amaranth and waterhemp throughout Ohio, there also does not seem to be a pattern in where resistant populations are located. Redroot pigweed populations that showed any level of resistance to any of the three sites of action (2, 9, 14) were located in nearly 50% of the counties that were surveyed in 2013 and 2014. In the 2014 experiment, nine of the populations screened for resistance were redroot pigweed that came from fields that received a rating of “1” for infestation levels, which represented individual plants in a field. Of these populations, 100% were resistant to the 1X and 4X dose of imazethapyr; approximately 50% were resistant to glyphosate at the 1X rate; and 100% were resistant

39 to fomesafen at the 1X rate with 33% being resistant at the 4X rate. These populations may have recently evolved herbicide-resistance, may be populations that have migrated from other areas, or much of the population was controlled by another herbicide; therefore, they have not yet completely infested the field. Controlling these populations before infestation is imperative, and it also important to monitor populations that survive at the end of the growing season even if they exist in smaller quantities relative to an infestation. Resistance in redroot pigweed is more widely distributed throughout the state than waterhemp and Palmer amaranth, likely due to its established presence throughout the state. Because redroot pigweed is abundant throughout all of Ohio, any populations that may be herbicide-resistant must be monitored to prevent the spread of resistant pigweed. Palmer amaranth and waterhemp are known to spread rapidly; therefore, the populations in Ohio must be managed to prevent any issues. The survey and greenhouse screen results allow growers to be aware of what issues they may be facing with

Amaranthus species, so they can attempt to prevent those issues. There are various proposed management tactics to combat and prevent herbicide-resistant weeds, and much work is being done to develop strategies for control of Amaranthus species. Among management recommendations is the use of preemergence herbicides with residual activity followed by a non-glyphosate POST herbicide treatment. Because some populations are resistant to PPO and ALS-inhibitors, these herbicides should also be avoided. Making herbicide applications before these target species are approximately seven centimeters is also important for effective control. Tillage and hand-weeding are other options for eliminating herbicide-resistant weed populations. New soybean technology with multiple herbicide resistance traits are also being developed and may be

40 useful in controlling and preventing further resistance evolution in Amaranthus species, as they will give producers a wider range of options for rotating sites of action.

41

Bibliography

Beckie, Hugh. “Herbicide-Resistant Weeds: Management Tactics and Practices.” Weed Technology 20(3): 793-814. 2006.

Heap, I. The International Survey of Herbicide Resistant Weeds. Online. Internet. Friday March 20, 2015. www.weedscience.org

Orson, J.H. “The Cost to the Farmer of Herbicide Resistance.” Weed Technology 13(3): 607-611. 1999.

Vencill, William K., Robert L. Nichols, Theodore M. Webster, John K. Soteres, Carol Mallory-Smith, Nilda R. Burgos, William G. Johnson, Marilyn R. McClelland. “Herbicide Resistance: Toward an Understanding of Resistance Development and the Impact of Herbicide Resistant Crops.” Weed Science 60(1): 2-30. 2012.

WSSA. “Herbicide Resistance and Herbicide Tolerance Definitions.” Weed Technology 12(4): 789. 1998.

42

Table 3.1 Response of 2013 Amaranthus spp. populations to herbicides in the greenhouse. R = resistant; S = susceptible; W = waterhemp; RR = redroot pigweed; P = Palmer. Glyphosate Fomesafen Imazethapyr Exp 1 Exp 2 Exp 1 Exp 2 Exp 1 Exp 2 1X 4X 1X 4X 1X 4X 1X 4X 1X 4X 1X 4X Sample Species

1 RR S S S S R S S S R R R R 2 RR S S S S S S S S S S S S 3 RR R R S S R S R S R R R S 4 RR R S S S R R S S R R R S 5 W R R R S S S S S R R R R 6 W R S R R S S S S R R R R 7 W R S R R R R

8 RR R S S S R R S S R S S S 9 RR R S R S R S

10 RR S S S S S S S S S S S S 11 RR S S S S S S S S S S R R 12 RR R S S S R S S S S S R S 13 RR S S S S R S S S S S R S 14 RR S S S S S S S S R R S S 15 RR S S S S R R S S R S S S 16 RR S S S S S S R S R S R R 17 W R S S S S S S S R S R R 18 RR S S S S S S R S R R R R 19 RR R S S S R S R S S S R R 20 RR S S S S S S S S S S R R 21 RR S S S S S S S S S S R S 22 RR S S R S S S

23 P

24 W R S S S S S R R R R S S 25 RR R S S S R R S S S S R S 26 P

27 RR R R S S R R R S R R R R 28 P S S S S S S R R R S R R 29 P S S S S S S S S R R R R 30 P

31 RR S S R R R S

Continued

43 Table 3.1: Continued 32 RR R S S S R S R S R R R R 33 RR S S R R S S

34 P S S S S R R

44 Table 3.2 Response of 2014 Amaranthus spp. populations to herbicides in the greenhouse. R = resistant; S = susceptible; W = waterhemp; RR = redroot pigweed; P = Palmer Glyphosate Fomesafen Imazethapyr

Sample Species 1X 4X 1X 4X 1X 4X 1 P S S R R R R 2 P

3 P

4 RR R S R S S S 5 P R S S S R R 6 P R S R S R R 7 P R R R S R R 8 W

9 P R S S S R R 10 RR S S R R R S 11 W R S S S R R 12 W R R R S R R 13 W R R S S R R 14 W R S S S R R 15 W R S R S R R 16 P R R S S R R 17 W R S R R S S 18 RR S S R R S S 19 W

20 W R R R R R R 21 W R S S S R R 22 W R S S S R R 23 W R R S S R R 24 RR S S R S R R 25 RR R S R S R R 26 P

27 W

28 W R S R R R R 29 W R S S S R R 30 W R R S S R R 31 RR R S R R R R 32 W R R S S R R 33 RR R S S S S S 34 RR R S R S S S 35 W

36 W

Continued 45 Table 3.2: Continued 37 W

38 RR R S R R R R 39 RR S S R R R R 40 RR S S R S R R 41 RR R S R S R R 42 RR S S R S R R 43 P S S R S R R 44 RR S S S S S S 45 W

46 RR S S R R R R 47 RR R R R S R R 48 W

49 RR R R R R R R 50 RR S S R S R R 51 RR R S R S R R 52 RR

46 Table 3.3 Percentage of 2013 Palmer, waterhemp, and redroot populations exhibiting resistance to site 2, 9, and 14 herbicides at 1X and 4X rates and populations exhibiting multiple resistance at 1X rates. Numbers in ( ) indicate number of populations. Experiment 1 Site 2 Site 9 Site 14 Site 2 + 9 Site 2 + 14 Site 9 + 14 Site 2 + 9 + 14 1X 4X 1X 4X 1X 4X Palmer (3) 100% 66% 0% 0% 0% 0% 0% (0) 0% (0) 0% (0) 0% (0) (3) (2) (0) (0) (0) (0) Waterhemp (5) 100% 80% 100% 20% 20% 20% 80% (4) 0% (0) 0% (0) 20% (1) (5) (4) (5) (1) (1) (1) Redroot (23) 50% 30% 40% 9% 65% 30% 0% (0) 13% (3) 13% (3) 26% (6) (12) (7) (9) (2) (15) (7) Experiment 2 Site 2 Site 9 Site 14 Site 2 + 9 Site 2 + 14 Site 9 + 14 Site 2 + 9 + 14 1X 4X 1X 4X 1X 4X Palmer (2) 100% 100% 0% 0% 50% 50% 0% (0) 50% (1) 0% (0) 0% (0) (2) (2) (0) (0) (1) (1) Waterhemp (4) 100% 75% 25% 25% 25% 25% 25% (1) 0% (0) 0% (0) 0% (0) (4) (3) (1) (1) (1) (1)

47 Redroot (19) 74% 42% 0% 0% 32% 0% 0% (0) 32% (6) 0% (0) 0% (0)

(14) (8) (0) (0) (6) (0)

47 Table 3.4 Percentage of 2014 Palmer, waterhemp, and redroot populations exhibiting resistance to site 2, 9, and 14 herbicides at 1X and 4X rates and populations exhibiting multiple resistance at 1X rates. Numbers in ( ) indicate number of populations. Site 2 Site 9 Site 14 Site 2 + 9 Site 2 + 14 Site 9 + 14 Site 2 + 9 + 14 1X 4X 1X 4X 1X 4X

Palmer (7) 100% 100% 71% 29% 57% 14% 43% (3) 14% (1) 14% (1) 29% (2) (7) (7) (5) (2) (4) (1) Waterhemp (14) 93% 93% 100% 43% 36% 21% 64% (9) 0% (0) 7% (1) 29% (4) (13) (13) (14) (6) (5) (3) Redroot (19) 74% 68% 53% 11% 89% 37% 0% (0) 57% (11) 16% (3) 37% (7) (14) (13) (10) (2) (12) (7)

48

48

Chapter 4: Control of Redroot Pigweed Utilizing Current and Prospective Herbicide

Technologies

Introduction

Since its introduction in 1974, glyphosate has become the most widely used herbicide in the world (Beckie, 2011). In 1996, glyphosate-resistant soybeans were introduced and have since been widely adopted. This gave farmers a simple and cost- effective method to weed control; therefore, the use of glyphosate for weed control in soybeans increased rapidly. From 1995, prior to the release of glyphosate-resistant soybean, to 2002, glyphosate use increased from 2.5 to 30 million kg/yr. Additionally, the number of applications in a single year increased, as well (Young, 2006). Initially, it was thought that glyphosate would contribute to the diversity of sites of action used in soybean weed control because it does not share the same site of action as any other herbicide; however, the obvious wide use has actually decreased the number of active ingredients farmers use for weed control in soybeans (Young, 2006). The overuse of glyphosate in crop production has resulted in glyphosate-resistant weeds. The first weed to be reported as glyphosate-resistant was horseweed (Conyza canadensis) in 2000

(Beckie, 2011). Presently, there are at least 14 weed species that are glyphosate-resistant in the United States (Heap, 2015). Among those species are weeds belonging to several different families, including the Amaranth family. Palmer amaranth (Amaranthus

49 palmeri) and tall waterhemp (Amaranthus rudis) both have multiple confirmed cases of glyphosate-resistance starting in 2005 (Heap, 2015).

With the growing number of herbicide-resistant weeds and the still frequent overuse of glyphosate, several suggestions have been made for preventing and managing herbicide-resistant weed populations. Riar et al. (2013) proposed several “best” management practices for herbicide-resistant weed populations. Those practices include:

1) Proper timing of herbicide application; 2) Planting into weed free fields; 3) Using multiple modes of action; 4) Using full herbicide rates; 5) Crop rotation; 6) Rotation of herbicide-resistant crop traits; 7) Use of cover crops; 8) Narrow crop row spacing; 9)

Altered planting date; 10) Tillage; and 11) Using sanitary equipment (Riar et al., 2013).

Hand-weeding is another option in extreme cases of herbicide-resistant weeds, which is usually a result of unsuccessful herbicide application. The rotation of herbicide-resistant crop traits in soybean will soon be easier to employ, as several new herbicide-resistant soybean varieties are in the approval stages. Many of these varieties contain more than one trait for herbicide resistance. This increases mode of action diversity making rotation easier.

In addition to the previously mentioned management tactics for herbicide- resistant weeds, an emphasis has also been put on the use of soil-applied herbicides with residual activity (Barnes & Oliver, 2004; Beckie, 2011; DeWerff et al., 2015; Whitaker et al., 2011). A trend of earlier soybean planting in the Midwest has been identified in recent years. With earlier planting, soybeans are subjected to a larger time frame for weed interference. Residual herbicides can be useful during this prolonged time period, so that postemergent applications can be delayed (DeWerff et al., 2015). Neve et al.’s

50 (2011) modeling work showed that the use of residual herbicides can delay development of herbicide resistance. Tank mixtures with residual herbicides applied preemergence or postemergence can increase control of herbicide resistant weeds. This provides alternative modes of action for resistance weed populations, as well as mode of action diversity to prevent resistance (Neve et al., 2011). Barnes and Oliver (2004) also showed that the use of residual herbicides in soybeans can improve weed control. Whitaker et al.

(2011) explored which residual herbicides would be most effective for controlling Palmer amaranth because previous research has showed that effective residual control is essential in management of glyphosate-resistant Palmer. They found that flumioxazin and pyrithiobac were the most effective residual herbicides applied preemergence. Residual herbicides can also be applied postemergence for season-long control (Whitaker et al.,

2011).

These studies approach control of redroot pigweed by using residual herbicides, combinations of herbicides currently labeled for use in soybean, and combinations of herbicides designed for new herbicide-resistant soybean technologies.

Objective

Develop management strategies for herbicide-resistant redroot pigweed using herbicides approved for use in currently available soybean varieties, and combinations of herbicides and prospective herbicide-resistant soybean technologies.

Materials & Methods

Two field studies were conducted in 2013 and 2014 at OARDC Western Ag

Research Station in South Charleston, Ohio. Methods for these studies were developed with other university investigators as part of a project funded by the United Soybean

51 Board to develop management solutions for glyphosate-resistant Amaranthus species in soybean production. The studies were initiated on May 22, 2013 and May 28, 2014. The redroot pigweed populations used for these studies were not resistant to any known herbicide sites of action. Redroot pigweed seed was spread in the experimental area both years. Other weeds were in the experimental area, such as giant ragweed (Ambrosia trifida), giant foxtail (Setaria faberi), ivyleaf morningglory (Ipomoea hederacea), velvetleaf (Abutilon theophrasti), common lambsquarters (Chenopodium album), Venice mallow (Hibiscus trionum) and barnyard grass (Echinochloa crus-galli).

The residual study evaluated the duration of control provided by a number of herbicides. Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 2 m wide by 7.6 m long. The experimental area was tilled prior to treatment application. Treatments were applied to bare soil using a

CO2 backpack sprayer with TeeJet AI XR 110015 nozzles calibrated to deliver 140 L/ha.

Treatments included dicamba, s-metolachlor, metribuzin, 2,4-D, isoxaflutole, pyroxasulfone, flumioxazin, and mesotrione applied individually and in combinations

(Table 4.1). Visual assessment of weed control was used to assess treatment effectiveness on redroot pigweed at 21 and 42 days after treatment. Assessment used a scale of 0 to 100, with 0 representing no control and 100 representing complete control.

Ratings were entered into Gylling Data Management ARM software. Data were analyzed as an ANOVA, and treatments were compared using the LSD (p = 0.05).

The herbicide programs study compared current soybean herbicide programs with those that could be used with prospective herbicide-resistant soybean varieties (Table

4.3). Methods were similar to those for the residual study except for application timings

52 and assessment timings. Application timings included: 1) PRE; 2) PRE followed by

POST 21 days later (EPOST); and 3) PRE followed by POST 42 days later (LPOST).

Control of redroot pigweed was assessed just prior to the first POST application, and then

21 and 42 days after each POST application, for a total of four assessments.

Results & Discussion

In the residual study, a number of treatments provided greater than 90% control at

21 DAT, but this often decreased considerably by the 42 DAT assessment.

Pyroxasulfone was the only herbicide that controlled 90% or more of the pigweed both years when applied alone. Other treatments controlling at least 90% of pigweed 21 DAT both years were: dicamba + s-metolachlor; dicamba + s-metolachlor + metribuzin; 2,4-D

+ s-metolachlor; isoxaflutole + s-metolachlor; isoxaflutole + s-metolachlor + metribuzin; pyroxasulfone; pyroxasulfone + flumioxazin; and mesotrione + s-metolachlor.

Additional treatments providing at least 84% control both years included metribuzin + either dicamba or isoxaflutole. Only two treatments provided at least 90% control 42

DAT. These were both combinations of three herbicides and included dicamba + s- metolachlor + metribuzin and isoxaflutole + s-metolachlor + metribuzin. These treatments can be used with prospective herbicide resistant soybean technology.

Dicamba + s-metolachlor + metribuzin could be used with soybeans resistant to dicamba, and isoxaflutole + s-metolachlor + metribuzin could be used with soybeans resistant to site 27 herbicides (HPPD inhibitors). Mesotrione + s-metolachlor controlled 86 and

93% of pigweed in 2013 and 2014, respectively, which was statistically similar to the previous three-way treatments. All three of these treatments resulted in less than 4 pigweed plants m2 42 DAT (Table 4.2). Metribuzin and 2,4-D were overall the least

53 effective herbicides, controlling no more than 73% of pigweed 21 DAT, and less than

50% at 42 DAT. When these two products were combined, they still only provided 78% and 67% control at 21 DAT in 2013 and 2014, respectively (Table 4.1). Population density 42 DAT ranged from 24 to 100 plants per m2 over both years for 2,4-D or metribuzin applied alone, and 15 to 21 plants per m2 in combination (Table 4.2).

In the herbicide programs trial, nearly all preemergence treatments provided 91% or above control at 21 DAT both years (Table 4.3) with no more than four pigweed plants per m2 (Table 4.4). Pyroxasulfone +flumioxazin followed by s-metolachlor + glufosinate

(EPOST) and dicamba + acetochlor followed by s-metolachlor + glyphosate + dicamba

(LPOST) were an exception to this, as they provided 100% control at 21 DAT in 2013 but only 75% control 21 DAT in 2014. Fomesafen + s-metolachlor + metribuzin was the only preemergence treatment that provided 90% or more control through 84 DAT in both years without a subsequent postemergence treatment (Table 4.3). Three other preemergence treatments controlled at least 80% of pigweed through 84 DAT, and included: pyroxasulfone + flumioxazin; s-metolachlor + metribuzin + isoxaflutole; dicamba + s-metolachlor . However, in several cases, when pyroxasulfone + flumioxazin was followed by either an EPOST or LPOST application, control improved.

Pyroxasulfone + flumioxazin followed by s-metolachlor + glyphosate (EPOST), which is a current herbicide program and pyroxasulfone + flumioxazin followed by s-metolachlor

+ glyphosate + dicamba (EPOST), which could be used with soybeans that are resistant to glyphosate and dicamba, provided 92% or more control through 84 DAT with one or less pigweed plants per m2, while pyroxasulfone + flumioxazin alone resulted in up to five pigweed plants per m2 (Table 4.3 and 4.4). Dicamba + acetochlor followed by s-

54 metolachlor + glyphosate + dicamba (EPOST) was the only other treatment with a preemergence followed by EPOST application that provided above 90% control at 84

DAT, and this treatment could also be used with soybeans resistant to glyphosate and dicamba. Several other treatments with a preemergence followed by EPOST application provided 83% or higher control, including pyroxasulfone + flumioxazin followed by s- metolachlor + glyphosate + 2,4-D (EPOST); pyroxasulfone + flumioxazin followed by s- metolachlor + glyphosate + 2,4-D + glufosinate (EPOST); s-metolachlor + metribuzin + isoxaflutole followed by glyphosate + fomesafen (EPOST); mesotrione + s-metolachlor + metribuzin followed by fomesafen + glyphosate (EPOST); and fomesafen + s- metolachlor + metribuzin followed by glyphosate + s-metolachlor + mesotrione (EPOST)

(Table 4.3).

Several treatments consisting of preemergence and LPOST applications provided

90% control through 84 DAT in both 2013 and 2014. All of these had a preemergence application of pyroxasulfone + flumioxazin followed by s-metolachlor + glyphosate; s- metolachlor + glyphosate + dicamba; s-metolachlor + glufosinate; s-metolachlor + glyphosate + 2,4-D; or s-metolachlor + glyphosate + 2,4-D + glufosinate 42 DAT. These treatments resulted in no more than 1.2 pigweed plants per m2 (Table 4.4). Two other pre followed by LPOST treatments provided upwards of 80% control at 84 DAT. These include mesotrione + s-metolachlor + metribuzin followed by fomesafen + glyphosate

(LPOST) and fomesafen + s-metolachlor + metribuzin followed by glyphosate + s- metolachlor + mesotrione (LPOST). For some treatments with the late post application, control from the preemergence herbicides occasionally decreased through the following

42 days, but increased after the postemergence application (Table 4.2) Examples of this

55 included dicamba + acetochlor followed by s-metolachlor + glyphosate + dicamba

(LPOST) and fomesafen + s-metolachlor + metribuzin followed by glyphosate + s- metolachlor + mesotrione + dicamba (LPOST) (Table 4.2).

The residual and herbicide programs studies showed that there are several options for managing redroot pigweed in soybeans with either herbicides currently labeled for use in soybeans or those that could be used with prospective herbicide-resistant soybean technologies. Some of the currently labeled active ingredients included pyroxasulfone, flumioxazin, metribuzin, and s-metolachlor. Glyphosate and glufosinate can also be used with glyphosate- and glufosinate-resistant soybean varieties, which are currently available and widely used. New varieties of herbicide-resistant soybeans are expected to include those that are resistant to HPPD inhibitors, 2,4-D + glufosinate; and dicamba, in conjunction with resistance to glyphosate and glufosinate. In the studies, there was no clear advantage between the currently available or prospeective herbicides. This is helpful for growers currently battling resistance or trying to prevent resistance, as these technologies combined provide several options for rotating sites of action. The prospective soybean technologies also allow many herbicides to be used either preemeregence or postemergence. Several herbicides were effective when applied preemergence and provided prolonged control, which would allow postemergence applications to be delayed to control later-emerging weeds. Current and future herbicide resistant soybean technologies can also be combined with other weed control methods to effectively control and prevent herbicide-resistant pigweed populations.

56

Bibliography

Barnes, Jeff W., Oliver, Lawrence R. “Preemergence Weed Control in Soybean with Cloransulam.” Weed Technology 18(4): 1077-1090. 2004.

Beckie, Hugh J. “Herbicide-resistant weed management: focus on glyphosate.” Pest Management Science 67: 1037-1048. 2011.

DeWerff, Ryan P., Conley, Shawn P., Colqquhoun, Jed B., Davis, Vince M. “Weed Control in Soybean as Influenced by Residual Herbicide Use and Glyphosate- Application Timing Following Different Planting Dates.” Weed Technology 29(1): 71-81. 2015.

Heap, I. The International Survey of Herbicide Resistant Weeds. Online. Internet. March 20, 2015. www.weedscience.org

Neve, Paul, Norsworthy, Jason K., Smith, Kenneth L., Zelaya, Ian A. “Modeling Glyphosate Resistance Management Strategies for Palmer Amaranth (Amaranthus palmeri) in Cotton.” Weed Technology 25(3): 335-343. 2011.

Riar, D., J. Norsworthy, L. Steckel, D. Stephenson, T. Eubank, J. Bond, and R. Scott. “Adoption of best management practices for herbicide-resistant weeds in midsouthern United States cotton, rice, and soybean.” Weed Technology 27(4): 788-797. 2013.

Whitaker, Jared R., York, Alan C., Jordan, David L., Culpepper, A. Stanley. “Palmer amaranth (Amaranthus palmeri) Control in Soybean with Glyphosate and Conventional Herbicide Systems.” Weed Technology 24(4): 403-410. 2010.

Whitaker, Jared R., York, Alan C., Jordan, David L., Culpepper, A. Stanley, Sosnoskie, Lynn M. “Residual Herbicides for Palmer Amaranth Control.” Journal of Cotton Science 15: 89-99. 2011.

Young, Bryan G. “Changes in Herbicide Use Patterns and Production Practices Resulting from Glyphosate-Resistant Crops.” Weed Technology 20(2): 301-307. 2006.

57

Table 4.1. Effect of soil-applied herbicides on residual control of redroot pigweed in the absence of a crop. Control Rate 21 DAT 42 DAT Treatment kg ai/ha % 2013 2014 2013 2014 dicamba 0.56 94 ab 71 de 50 fgh 17 h dicamba 1.12 95 ab 78 cd 60 def 51 def s-metolachlor 1.07 87 bc 78 cd 45 fgh 52 de metribuzin 0.78 65 ef 54 fg 32 hi 37 efg dicamba 1.12 99 a 97 a 76 abcd 80 ab s-metolachlor 1.07 dicamba 1.12 98 ab 88 abc 75 bcd 73 bc metribuzin 0.78 2, 4-D 0.56 55 f 45 g 30 i 43 def 2, 4-D 1.12 73 de 55 fg 40 ghi 22 gh 2,4-D 1.12 90 abc 93 ab 60 efg 56 cde s-metolachlor 1.07 2,4-D 1.12 78 cd 67 de 53 efg 42 fg metribuzin 0.78 dicamba 1.12 100 a 100 a 90 ab 96 a s-metolachlor 1.07 metribuzin 0.78 isoxaflutole 0.1 92 ab 65 ef 71 cde 32 fgh isoxaflutole 0.1 99 ab 91 ab 77 abcd 73 bc s-metolachlor 1.07 isoxaflutole 0.1 97 ab 84 bc 80 abc 58 cd metribuzin 0.78 isoxaflutole 0.1 100 a 100 a 93 a 93 a s-metolachlor 1.07 metribuzin 0.78 pyroxasulfone 0.44 93 ab 90 ab 52 fg 91 ab pyroxasulfone 0.09 97 ab 93 ab 78 abc 81 ab flumioxazin 0.07 mesotrione 0.18 100 a 99 a 86 abc 93 a s-metolachlor 1.21 UTC a Means within a column are not significantly different based on LSD p = 0.05

58

Table 4.2 Effect of soil-applied herbicides on population density of redroot pigweed in the absence of a crop Treatment Rate Density 42 DAT kg ai/ha plants per 1 m2 2013 2014 dicamba 0.56 16 bcde 78 b dicamba 1.12 19 abcde 22 cdef s-metolachlor 1.07 21 abcd 37 cd metribuzin 0.78 24 abc 42 c dicamba 1.12 10 def 8.6 def s-metolachlor 1.07 dicamba 1.12 5 fgh 7.6 def metribuzin 0.78 2, 4-D 0.56 36 a 104 b 2, 4-D 1.12 23 abc 84 b 2,4-D 1.12 15 cde 32 cde s-metolachlor 1.07 2,4-D 1.12 15 cde 21 cdef metribuzin 0.78 dicamba 1.12 3.2 ghi 1 f s-metolachlor 1.07 metribuzin 0.78 isoxaflutole 0.1 9.6 ef 85 b isoxaflutole 0.1 3.6 gh 10 def s-metolachlor 1.07 isoxaflutole 0.1 6.4 fg 5.6 ef metribuzin 0.78 isoxaflutole 0.1 0 j 1.6 ef s-metolachlor 1.07 metribuzin 0.78 pyroxasulfone 0.44 5.2 fgh 2 ef pyroxasulfone 0.09 1.8 hij 3.6 ef flumioxazin 0.07 mesotrione 0.18 0.6 ij 3.6 ef s-metolachlor 1.21 UTC 33 ab 139 a a Means within a column are not significantly different based on LSD p = 0.05

59 Table 4.3 Effect of soil- and foliar-applied herbicides on control of redroot pigweed in the absence of a crop Control % Treatment Rate Timing 2013 2014 kg ai/ha 21 DAT 42 DAT 63 DAT 84 DAT 21 DAT 42 DAT 63 DAT 84 DAT UTC pyroxasulfone 0.09 flumioxazin 0.07 PRE 99 a 90 a 86 bc 94 a 100 a 96 a 99 a 87 bcdefg s-metolachlor 1.07 metribuzin 0.42 PRE 100 a 99 a 99 ab 99 a 100 a 99 a 81 c 82 efh isoxaflutole 0.1 dicamba 1.12 acetochlor 2.3 PRE 100 a 76 bc 73 c 84 a 100 a 100 a 92 b 75 gh mesotrione 0.18 s-metolachlor 1.21 PRE 100 a 100 a 100 a 99 a 99 a 96 a 86 c 84 defg metribuzin 0.28

60 fomesafen 0.26

s-metolachlor 1.87 PRE 98 a 96 a 91 ab 97 a 100 a 92 a 90 b 93 abcde metribuzin 0.28 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 99 a 100 a 100 a 99 a 100 a 99 a 99 a 92 abcde glyphosate 0.86 DAT pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 99 a 100 a 100 a 100 a 100 a 100 a 95 ab 94 abcde glyphosate 0.86 DAT dicamba 0.56

Continued

60 Table 4.3: Continued dicamba 1.12 PRE acetochlor 2.3 s-metolachlor 1.07 21 100 a 100 a 95 ab 97 a 99 a 99 a 96 ab 90 abcdef glyphosate 0.86 DAT dicamba 0.56 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 100 a 100 a 100 a 100 a 75 b 99 a 94 ab 78 f glufosinate 0.59 DAT pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 100 a 100 a 100 a 100 a 100 a 100 a 99 a 85 cdefg glyphosate 0.86 DAT 2,4-D 1.12 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 99 a 100 a 97 ab 100 a 100 a 100 a 97 ab 85 cdefg glyphosate 0.86 DAT 61 2,4-D 0.56 glufosinate 0.59 s-metolachlor 1.07 PRE metribuzin 0.42 isoxaflutole 0.1 100 a 100 a 98 ab 99 a 99 a 100 a 91 b 83 defg glyphosate 0.86 21 fomesafen 0.26 DAT mesotrione 0.18 PRE s-metolachlor 1.87 metribuzin 0.28 fomesafen 0.32 21 100 a 99 a 97 ab 100 a 100 a 100 a 98 ab 83 defg glyphosate 1.29 DAT MSO 1% v/v

Continued

61

Table 4.3: Continued fomesafen 0.26 PRE s-metolachlor 1.21 metribuzin 0.28 glyphosate 1.05 21 98 a 100 a 100 a 99 a 98 a 100 a 98 ab 85 cdefg s-metolachlor 1.05 DAT mesotrione 0.1 Induce 0.25% v/v fomesafen 0.26 PRE s-metolachlor 1.87 metribuzin 0.28 glyphosate 1.05 21 s-metolachlor 1.05 DAT 99 a 100 a 100 a 99 a 91 a 100 a 93 b 70 h mesotrione 0.1 dicamba 0.56 Induce 0.25% v/v pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 100 a 99 a 99 ab 95 a 100 a 100 a 98 ab 95 abc 6

2 glyphosate 0.86 DAT pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 99 a 99 a 100 a 99 a 100 a 100 a 100 a 98 ab glyphosate 0.86 DAT dicamba 0.56 dicamba 1.12 PRE acetochlor 2.3 s-metolachlor 1.07 42 100 a 89 ab 100 a 99 a 75 b 66 b 98 ab 98 ab glyphosate 0.86 DAT dicamba 0.56 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 99 a 99 a 96 ab 99 a 99 a 100 a 97 ab 94 abcde glufosinate 0.59 DAT

Continued 62 Table 4.3: Continued pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 100 a 99 a 100 a 98 a 100 a 100 a 100 a 100 a glyphosate 0.86 DAT 2,4-D 1.12 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 glyphosate 0.86 DAT 98 a 94 a 100 a 98 a 100 a 100 a 100 a 98 abc 2,4-D 0.56 glufosinate 0.59 s-metolachlor 1.07 PRE metribuzin 0.42 isoxaflutole 0.1 99 a 88 ab 88 ab 99 a 100 a 97 a 98 ab 91 abcde glyphosate 0.86 42 fomesafen 0.26 DAT mesotrione 0.18 PRE s-metolachlor 1.87

6 metribuzin 0.28 3 100 a 100 a 100 a 100 a 98 a 98 a 98 ab 87 bcdefg fomesafen 0.32 42 glyphosate 1.29 DAT MSO 1% v/v fomesafen 0.26 PRE s-metolachlor 1.21 metribuzin 0.28 glyphosate 1.05 42 92 a 86 ab 98 ab 97 a 100 a 91 a 98 ab 97 abc s-metolachlor 1.05 DAT mesotrione 0.1 Induce 0.25% v/v

Continued

63 Table 4.3: Continued fomesafen 0.26 PRE s-metolachlor 1.21 metribuzin 0.28 glyphosate 1.05 42 s-metolachlor 1.05 DAT 96 a 72 c 100 a 88 a 100 a 91 a 100 a 97 abc mesotrione 0.1 dicamba 0.56 Induce 0.25% v/v

a Means within a column are not significantly different based on LSD p = 0.05 6 4

64 Table 4.4 Effect of soil- and foliar-applied herbicides on population density of redroot pigweed in the absence of a crop Density plants per 1 m2 21 days after PRE 21 days after EPOST 21 days after LPOST Treatment Rate Timing 2013 2014 2013 2014 2013 2014 kg ai/ha UTC 0.6 bc 186 a 0.8 abcde 105 a 14 a 61 a pyroxasulfone 0.09 PRE flumioxazin 0.07 0 c 0 c 0.2 cde 0.5 b 5 c 0.5 bc s-metolachlor 1.07 PRE metribuzin 0.42 0 c 0 c 0.2 cde 0.2 b 10 ab 2.5 bc isoxaflutole 0.1 dicamba 1.12 PRE 0 c 0 c 1 abcde 0.5 b 6 bc 2.5 bc acetochlor 2.3 mesotrione 0.18 PRE s-metolachlor 1.21 0 c 0 c 0.8 bcde 0.7 b 5 c 1.5 bc metribuzin 0.28

6 fomesafen 0.26 PRE 5

s-metolachlor 1.87 0 c 0 c 0.6 bcde 2.2 b 4 cd 1 bc metribuzin 0.28 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 DAT 0 c 0 c 0 e 0 b 0 d 1 bc glyphosate 0.86 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 DAT 0 c 0 c 0 e 0 b 0 d 0.5 bc glyphosate 0.86 dicamba 0.56

Continued

65 Table 4.4: Continued dicamba 1.12 PRE acetochlor 2.3 s-metolachlor 1.07 21 DAT 0.2 bc 0 c 0 e 0 b 0 d 1 bc glyphosate 0.86 dicamba 0.56 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 DAT 0 c 2 bc 0 e 1 b 0 d 1.2 bc glufosinate 0.59 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 DAT 0 c 0 c 0 e 0 b 0 d 0.7 bc glyphosate 0.86 2,4-D 1.12 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 21 DAT glyphosate 0.86 0 c 0 c 0 e 0 b 0 d 1 bc

6 2,4-D 0.56

6 glufosinate 0.59

s-metolachlor 1.07 PRE metribuzin 0.42 isoxaflutole 0.1 0 c 0 c 0 e 0 b 0 d 0.7 bc glyphosate 0.86 21 DAT fomesafen 0.26 mesotrione 0.18 PRE s-metolachlor 1.87 metribuzin 0.28 fomesafen 0.32 21 DAT 0.2 bc 0 c 0 e 0 b 0 d 1 bc glyphosate 1.29 MSO 1% v/v

Continued

66 Table 4.4: Continued fomesafen 0.26 PRE s-metolachlor 1.21 metribuzin 0.28 glyphosate 1.05 21 DAT 0 c 2 bc 0 e 0 b 0 d 0.5 bc s-metolachlor 1.05 mesotrione 0.1 Induce 0.25% v/v fomesafen 0.26 PRE s-metolachlor 1.87 metribuzin 0.28 glyphosate 1.05 21 DAT 0 c 4 bc 0 e 0 b 0 d 1 bc s-metolachlor 1.05 mesotrione 0.1 dicamba 0.56 Induce 0.25% v/v pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 DAT 0.2 bc 0 c 0 e 0 b 0 d 0 c glyphosate 0.86 pyroxasulfone 0.09 PRE 67 flumioxazin 0.07

s-metolachlor 1.07 42 DAT 0 c 2 bc 0.2 de 0 b 0 d 0 c glyphosate 0.86 dicamba 0.56 dicamba 1.12 PRE acetochlor 2.3 s-metolachlor 1.07 42 DAT 1 ab 42 b 1.2 abcde 2.5 b 0 d 7.5 b glyphosate 0.86 dicamba 0.56 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 DAT 0.8 bc 2 bc 1.4 abcd 0.5 b 0 d 0 c glufosinate 0.59

Continued

67 Table 4.4: Continued pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 DAT 0 c 2 bc 0.2 cde 1.7 b 0 d 0.5 bc glyphosate 0.86 2,4-D 1.12 pyroxasulfone 0.09 PRE flumioxazin 0.07 s-metolachlor 1.07 42 DAT glyphosate 0.86 0.2 bc 2 bc 0.2 cde 0.5 b 0 d 0 c 2,4-D 0.56 glufosinate 0.59 s-metolachlor 1.07 PRE metribuzin 0.42 isoxaflutole 0.1 1.4 ab 0 c 2.2 abc 0.7 b 0 d 0.5 bc glyphosate 0.86 42 DAT fomesafen 0.26 mesotrione 0.18 PRE s-metolachlor 1.87

68 metribuzin 0.28 0 c 0 c 0 e 0 b 0 d 2.5 bc fomesafen 0.32 42 DAT glyphosate 1.29 MSO 1% v/v fomesafen 0.26 PRE s-metolachlor 1.21 metribuzin 0.28 glyphosate 1.05 42 DAT 3.4 a 2 bc 4 a 0.7 b 0 d 0 c s-metolachlor 1.05 mesotrione 0.1 Induce 0.25% v/v

Continued

68 Table 4.4: Continued fomesafen 0.26 PRE s-metolachlor 1.21 metribuzin 0.28 glyphosate 1.05 42 DAT s-metolachlor 1.05 0.8 bc 4 bc 2.8 ab 3 b 0 d 0 c mesotrione 0.1 dicamba 0.56 Induce 0.25% v/v

a Means within a column are not significantly different based on LSD p = 0.05 69

69

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