Chrysodeixis Includens (Walker), Thresholds In

Refinement and validation of soybean looper, Chrysodeixis includens (Walker), thresholds in

Mississippi soybean Glycine max (L.) Merr.

By TITLE PAGE Mary Kathryn Huff

A Thesis Submitted to the Faculty of Mississippi State University in Partial Fulfillment of the Requirements for the Degree of Master of Science in Agricultural Life Sciences in the Department of Biochemistry, Molecular Biology, Entomology, and Plant Pathology

Mississippi State, Mississippi

May 2020

2020

Refinement and validation of soybean looper (Chrysodeixis includens) (Walker) thresholds in

Mississippi soybeans (Glycine max (L.) Merr.)

By APPROVAL PAGE Mary Kathryn Huff

Approved:

______Donald R. Cook (Major Professor)

______Angus L. Catchot Jr. (Co-Major Professor)

______Jeffrey Gore (Committee Member)

______Fred R. Musser (Committee Member)

______Jon Trenton Irby (Committee Member)

______Kenneth O. Willeford (Graduate Coordinator)

______George M. Hopper Dean College of Agriculture and Life Sciences

Name: Mary Kathryn Huff ABSTRACT Date of Degree: May 1, 2020

Institution: Mississippi State University

Major Field: Agricultural Life Sciences

Major Professors: Donald R. Cook and Angus L. Catchot Jr.

Title of Study: Refinement and validation of soybean looper (Chrysodeixis includens) (Walker) thresholds in Mississippi soybeans (Glycine max (L.) Merr.)

Pages in Study: 88

Candidate for Degree of Master of Science

Experiments were conducted to refine and validate the soybean looper, Chrysodeixis includens (Walker) threshold in Mississippi soybeans, Glycine max. (L.) Merr. Equivalencies between sweep net and drop cloth sampling methods were evaluated, overall the sweep net was more effective at capturing soybean looper larvae greater than third instar and larvae less than or equal to third instar when compared to the drop cloth method. Feeding studies were conducted to determine the feeding rates of each instar, results showed that fifth instar larvae consume the greatest amount of leaf tissue, and larvae greater than or equal to third instar consume more leaf tissue than larvae less than third instar. Observed and expected/predicted defoliation rates were calculated. Observed defoliation was lower than predicted defoliation possibly due to field environmental conditions. Using data in combination with published data from other studies, three prediction models were created for the R3, R5, and R6 growth stage.

DEDICATION

This thesis is dedicated to my late great-grandmother, Gwendolyn King. The memory of her unconditional love and her unshakable strength have always driven me and will continue to do so all the days of my life. I could spend the rest of my days describing the impact she made on my life, but her impact and her love are best summed up by Albus Dumbledore in the Harry

Potter series by J.K. Rowling when he said, “…love as powerful as [her’s] for you leaves its own mark. Not a scar, no visible sign… to have been loved so deeply, even though the person who loved us is gone, will give us some protection forever.” And for her love, guidance, and presence

I will be forever grateful. Her greatest wish in life was to see me graduate high school and although she never got to see that wish come to fruition, I can only hope that she is looking down on me now and that she is proud.

This thesis is also dedicated to my family. To my parents, Watt and Susie Huff, I could not be more grateful for the two of you. Even though you still don’t know exactly what I’m getting a degree in, and for a time thought I was getting a master’s degree in soybeans, your support has never wavered. I am so incredibly thankful for the values you have instilled in me and for your unconditional love and support. To my cousins, Lauren, Sarah, and Rachel; the constant laughs and honest advice from the three of you has been the greatest support system I could ever ask for.

ACKNOWLEDGEMENTS

I would first like to acknowledge my advisors and committee members. I cannot thank you enough for giving me this opportunity to learn and to pursue a Master’s degree. The knowledge and advice you all have given me will never be taken for granted, nor will the patience and guidance you have shown me.

Next I would like to acknowledge my fellow graduate students. To Joel Moor and Read

Kelly, thank you for helping me navigate my way through classes and writing papers, I could not have passed without you. To Will Hardman, Sara Barrett, Russ Godbold, and Cade Francis, thank you for always being there when I needed a helping a hand. When I look back on my time in grad school I will always remember the laughs and long conversations, you all have been (and hopefully will continue to be) the very best of friends.

To the full time staff and summer workers at DREC: Aaron Leininger, Meg Cutts, Neil

Wright, Hauff Carpenter, Harper Horton, Rainer Hodges, Duncan Henson, Jamarion Merrill,

Trey Freeland, Hunter Price, Jasmine Warren, and countless others that jumped in and lent a hand. I could not have built cages, pulled leaves off of hundreds of plants, weighed larvae, or sampled fields without you. No amount of lunches bought or thank you’s said will be enough to show my gratitude to all of you.

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TABLE OF CONTENTS

DEDICATION ...... ii

ACKNOWLEDGEMENTS ...... iii

TABLE OF CONTENTS ...... iv

LIST OF TABLES ...... vi

LIST OF FIGURES ...... viii

CHAPTER

I. LITERATURE REVIEW ...... 1

Soybeans ...... 1 Soybean Defoliation ...... 3 Sampling Larvae on Soybeans ...... 4 Soybean Looper ...... 6 Soybean Thresholds ...... 11 References Cited ...... 15

II. EVALUATION OF DROP CLOTH AND SWEEPNET SAMPLING METHODS FOR SOYBEAN LOOPER INFESTING SOYBEAN IN MISSISSIPPI ...... 22

Abstract ...... 22 Introduction ...... 22 Materials and Methods ...... 25 Results ...... 27 Discussion ...... 28 References Cited ...... 38

III. EVALUATION OF SOYBEAN LOOPER LARVAL CONSUMPTION RATES ...... 40

Abstract ...... 40 Introduction ...... 41 Materials and Methods ...... 42 Results ...... 44 iv

Discussion ...... 46 References Cited ...... 56

IV. REFINEMENT AND VALIDATION OF SOYBEAN LOOPER LARVAL ECONOMIC THRESHOLD FOR SOYBEANS IN MISSISSIPPI ...... 57

Abstract ...... 57 Introduction ...... 58 Materials and Methods ...... 59 Results and Discussion ...... 61 References Cited ...... 82

V. SUMMARY OF STUDIES ...... 85

References Cited ...... 88

LIST OF TABLES

Table 1.1 Defoliation and sampling thresholds for soybean looper larvae in soybean producing states...... 14

Table 2.1 Environmental data, row spacing, and crop height for sampling sites during 2017 and 2018...... 31

Table 3.1 Date, location, number of individuals collected, and mortality rates for 2017 and 2018 collections ...... 49

Table 3.2 Soybean looper larval feeding rates from this study compared with results from (Reid and Green 1973, Kogan and Cope 1974, Boldt et al. 1975, Trichillo and Mack 1989)...... 50

Table 3.3 Conservative, moderate, and aggressive feeding rate and insect equivalent models for soybean looper larvae feeding on soybean leaves ...... 51

Table 4.1 Regression analysis equation for the conservative, moderate, and aggressive feeding models ...... 64

Table 4.2 Economic injury levels at R3 and R5 for defoliation based on the yield loss equation for R3 growth stage soybean...... 64

Table 4.3 Economic injury levels for soybean looper infesting soybean at R3 based on defoliation converted to soybean looper insect equivalents for three feeding models...... 65

Table 4.4 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the conservative feeding model at R3 growth stage ...... 66

Table 4.5 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the moderate feeding model at R3 growth stage...... 67

Table 4.6 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the aggressive feeding model at R3 growth stage...... 68

Table 4.7 Economic injury levels at R3 and R5 for defoliation based on the yield loss equation for R5 growth stage...... 69

Table 4.8 Economic injury levels for soybean looper infesting soybean at R5 based on defoliation converted to soybean looper insect equivalents for three feeding models...... 70

Table 4.9 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the conservative feeding model for the R5 growth stage...... 71

Table 4.10 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the moderate feeding model at the R5 growth stage...... 72

Table 4.11 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the aggressive feeding model at the R5 growth stage...... 73

Table 4.12 Economic injury level at R6 for defoliation based on the yield loss equation at R6 growth stage ...... 74

Table 4.13 Economic injury levels for soybean looper infesting soybean at R6 based on defoliation converted to soybean looper insect equivalents for three feeding models...... 75

Table 4.14 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the conservative feeding model at the R6 growth stage...... 76

Table 4.15 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the moderate feeding model at the R6 growth stage...... 77

Table 4.16 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the aggressive feeding model at the R6 growth stage...... 78

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LIST OF FIGURES

Figure 2.1 Relationship between numbers of soybean looper larvae less than third instar captured using drop cloth and sweep net sampling...... 32

Figure 2.2 Relationship between numbers of third instar soybean looper larvae captured using sweep net and drop cloth sampling...... 33

Figure 2.3 Relationship between numbers of fourth instar soybean looper larvae captured using drop cloth and sweep net sampling...... 34

Figure 2.4 Relationship between numbers of fifth instar soybean looper larvae captured using sweep net and drop cloth sampling...... 35

Figure 2.5 Relationship between numbers of soybean looper larvae greater than or equal to third instar captured using drop cloth and sweep net sampling...... 36

Figure 2.6 Relationship between total number of soybean looper larvae captured using drop cloth and sweep net sampling methods...... 37

Figure 3.1 Leaf area consumption (cm2) (±SE) of first through fifth instar soybean looper larvae during 2019. Leaves were obtained from greenhouse grown MG IV soybeans. Larvae were obtained from Benzon Research...... 52

Figure 3.2 Mean leaf consumption (cm2) (±SE) per day of first through fifth instar soybean looper larvae during 2019. Leaves were obtained from greenhouse grown MG IV soybeans. Larvae were obtained from Benzon Research...... 53

Figure 3.3 Leaf tissue consumption by weight (mg) (±SE) for first through fifth instar soybean looper larvae during 2019. Leaves were obtained from greenhouse grown MG IV soybeans. Larvae were obtained from Benzon Research...... 54

Figure 3.4 Weight gain (mg) (±SE) of soybean looper larvae during the second, third, fourth, and fifth instar of development during 2019. Larvae were obtained from Benzon Research and were fed leaves obtained from greenhouse grown MG IV soybeans...... 55

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Figure 4.1 Relationship between observed percent defoliation and number of soybean looper insect equivalents calculated based on the conservative model of over a 10-day period...... 79

Figure 4.2 Relationship between observed percent defoliation and number of soybean looper insect equivalents calculated based on the moderate model of feeding over a 10-day period...... 80

Figure 4.3 Relationship between observed percent defoliation and number of soybean looper insect equivalents calculated based on the aggressive model of feeding over a 10-day period...... 81

CHAPTER I

LITERATURE REVIEW

Soybeans

In the last decade, annual soybean, Glycine max (L.) Merr. production in Mississippi averaged ca. 800,000 ha. Average yields of soybean in Mississippi was 2,959 kg per ha, and a value of $0.30 to $0.38 per kg. (USDA NASS). Since the late 1990s, soybean production has shifted to an Early Season Planting System (ESPS) to reduce risks from drought stress, improve profitability, and avoid late season insect pest infestations (McPherson et al. 2001). The ESPS involves seedbed preparation in the fall, applying a pre-plant herbicide to kill winter/spring weeds, and planting early-maturing varieties into a stale seedbed in April (Heatherly and Spurlock 1999).

Soybeans are photosensitive, and plant reproduction is directly related to the amount of light received during daylight hours (Ashlock et al. 2014). As a result of this sensitivity, varieties in

North America are adapted for growth within 180 to 300 km wide latitudinal bands (Tanner and

Hume 1978). Cultivars have been bred into 13 maturity groups ranging from MG000 (fastest maturing) to MG X (slowest maturing) (Brown et al. 2017). Each of these varieties are more adapted to certain latitudinal bands based on day length. The early maturing varieties (000 – IV) typically have an indeterminate type of growth, while the later maturing varieties (V-X) have a determinate type of growth. Indeterminate maturity groups continue new vegetative growth after flowering begins and continue vegetative growth through the early to mid-reproductive stages.

Determinate maturity groups cease vegetative growth soon after reproductive stages begin;

reproductive development occurs uniformly along the main stem of the plant compared to indeterminate soybeans. Using mid to early-maturing varieties in Mississippi (commonly MG IV and V) decreases the amount of time needed from emergence to flowering and shortens the growing period (McPherson et al. 2001).

Prior to the 1970s, there was no standard system for describing the growth stages of soybeans. Fehr et al. (1971) devised a method using the number of nodes along the main stem of the plant during the vegetative stages and the appearance of flowers, pods, and seeds to describe the reproductive stages. Following the emergence of the radicle, the hypocotyl will pull the cotyledons out of the soil and the plant will be at stage VE, or emergence, once the cotyledons and unifoliates have emerged and fully expanded, stage VC is reached (Fehr et al. 1971, Koger et al. 2014). As the plant develops, trifoliates begin to emerge, once the leaves have unrolled and the individual leaflets do not touch, stage V1 is achieved, each subsequent trifoliate emergence marks a new growth stage (Vn). From VC to V5 a new stage occurs every 5 to 7 days; from V5 to the beginning of reproduction, a new growth stage will occur every 3 to 5 days under optimal conditions (Koger et al. 2014). For indeterminate varietals, the occurrence of any open flower on any node on the main stem, the reproductive growth stages begin (R1 – beginning bloom), this stage typically begins at the third to sixth node on the plant and lasts 3 to 4 weeks (Koger et al.

2014). As flowering continues to occur, an open flower on one of the two uppermost main-stem nodes marks full bloom (R2) (Fehr et al. 1971). The presence of any pod 0.5 cm (3/16 inch) or longer on one of the four uppermost nodes of the main stem signifies the beginning pod set (R3), once one pod at one of the four uppermost nodes on the main stem lengthens to 1.905 cm (3/4 inch) long, full pod set (R4) has occurred (Fehr et al. 1971). The beginning of seed formation

(R5), is primarily marked by the occurrence of a seed 0.3175 cm (1/8 inch) long in one of the

pods on one of the uppermost four nodes (Fehr et al. 1971). At R5, the soybean plant has reached its maximum height, node number, leaf area, and nitrogen fixation capacity (Koger et al. 2014).

Once a pod on one of the four uppermost nodes contains a green seed that fills the pod capacity, the plant is at full seed (R6). As the plant starts shedding leaves and one pod with mature color occurs anywhere on the plant, maturity has begun (R7). Once 95% of pods have reached their mature color, the soybean plant is considered to be at full maturity (R8) (Fehr et al. 1971).

Soybean Defoliation

Defoliation is considered the most common and most visible type of injury to soybeans.

Defoliation primarily refers to all types of injury that results in a reduction of the total functioning leaf surface of a plant (Kogan and Turnipseed 1980). According to Herbert et al.

(1992), soybean defoliation typically occurs in two patterns: 1) the gradual removal of leaf tissue that typically does not exceed 25% of the total leaf area occurring before reproductive growth stages and 2) more severe defoliation of 50 to 100% by pests that occurs within a two-week period, most commonly during the R5 to R6 growth stages. Yield losses resulting from defoliation are primarily due to reduced light interception (Browde et al. 1994) leading to decreased photosynthetic activity, removal of stored carbohydrates and nitrogen, loss of storage material, shortening of the effective pod set and filling growth stages, and an increase in water loss (Pickle and Caviness 1984, Bidwell 1979). The amount of yield reduction from defoliation of soybeans is a function of the stage of plant development when the injury occurs (Fehr et al.

1971). Prior to flowering, soybeans can withstand 33 – 53% defoliation with little yield loss

(Kalton et al. 1945, Camery and Weber 1953, Turnipseed 1972, Todd and Morgan 1971, Thrash

2018). Pickle and Caviness (1984) reported that less than 50% defoliation did not reduce yields, but additional studies report that soybeans can tolerate up to 75% defoliation before yield loss 3

(Fuellman 1944, Kalton et al. 1945). During pod formation (R3) to the beginning of seed formation (R5), soybeans are most sensitive to yield loss from defoliation (Talekar and Lee

1988, Fuellman 1944, Kalton et al. 1945, McAlister and Krober 1958, Turnipseed 1972). At these stages, prior reports noted that 33 – 50% defoliation events significantly reduced yield

(Talekar and Lee 1988, Thomas et al. 1974). Defoliation of 10, 25, or 50 % during two consecutive growth stages or 25 and 50% during three consecutive growth stages significantly reduced yield (Talekar and Lee 1988). However, much of this research was done prior to the adoption of the modern high yielding varieties and may not be consistent with more recent research. Current research has established thresholds of 35% defoliation prior to flowering and

20% defoliation during reproductive stages without any impact on yield (Owen et al. 2013).

Additionally, defoliation events occurring at different growth stages are independent of each other and that continuous sub-threshold levels of defoliation do not compound upon each other to cause significant yield losses (Thrash 2018). After pod fill has completed (R6.5), effects from defoliation are not considered significant, even at very high levels (Malone et al. 2002, Kalton et al. 1945, Turnipseed 1972).

Sampling Larvae on Soybeans

In the past century, four primary methods of entomological sampling in soybeans have been identified: direct observations, ground or drop cloth, sweep net, and vacuum net (Kogan and Turnipseed 1980). The two primary methods of sampling soybean are the sweep net and drop cloth. Of these two methods, the drop cloth is sometimes considered absolute population estimator for many species and the sweep net is considered a relative sampling method (Kogan and Pitre 1980). For most pest management purposes, obtaining relative population estimates is usually sufficient in situations where rapid classification into decision categories is necessary 4

(e.g., spray or do not spray) (Ruesink and Kogan 1975). The drop cloth is ideal for sampling slow moving arthropods that are easily dislodged from plants. This method is limited when plants are small and becomes inefficient when plants begin to senesce and shed their leaves

(Kogan and Pitre 1980). In soybeans, this sampling technique is widely recommended for scouting defoliators and pod feeders, however the drop cloth method varies in effectiveness depending upon the species composition and the plant growth stage (Strayer and Green 1974,

Barnes et al. 1974, Kogan et al. 1977, Marston et al. 1979). Recovery efficiency of larvae may be related to larval behavior. For example, soybean looper, Chrysodeixis includens (Walker) larvae stay motionless when disturbed (Marston et al. 1979). Sampler variability can be counteracted by ensuring that different surveyors shake the plants with similar amounts of vigor (Kogan and Pitre

1980, Marston et al. 1979). The sweep net is more widely used than the drop cloth, because it can capture more insects from vegetation per man hour without an increased cost for equipment and damage to the crop. Despite its popularity, there has been difficulties in standardizing the sweep net method for accurate population studies (Ruesink and Kogan 1975). Much of the variation of the sweep net can be attributed to the following factors: temperature, humidity, wind velocity, position of the sun, plant size, density of the canopy, and pubescence of leaves and stems. Temperature can influence the metabolic rates of insects which affects the speed of escape reactions. Humidity, combined with temperature, can affect the microclimate within the canopy causing changes in the individual’s position on the plant. Wind velocity affects the location of certain species that may take shelter deeper into the canopy in heavy winds. Position of the sun can cause shadows to be cast which can cause certain species to react and escape. The plant size affects sweeping as smaller plants are too fragile to sweep and the larger plants can only be partly sampled. As canopy density increases, there will be increased resistance to sweep net

movements which limits efficiency and larger plants will offer more protection for insects.

Finally, the pubescence of leaves and stems can lessen the impact of the sweep net strokes or provide additional shelter. Despite all of these environmental factors, the majority of the variation comes from human factors such as: height of sweeping, rapidity and length of the sweep net stroke, and the sweeping method (DeLong 1932). Studebaker et al. (1991) indicates that neither the drop cloth nor the sweep net can be considered absolute methods for sampling soybean looper in soybeans. However, both methods are related to absolute densities adequately enough to be useful in sampling programs. (Studebaker et al. 1991). Sampling and population estimation variation can result from samplers’ experience in counting mobile species, the ability to rapidly differentiate between species, and sampler fatigue which can result in decreased accuracy after several hours of sustained work (Marston et al. 1979). Row spacing may also affect the efficacy of both sampling methods. Wide row plantings are easily sampled with either a drop cloth or sweep net, whereas narrow row spacing makes sampling with a drop cloth virtually impossible (Studebaker et al. 1991, Pedigo et al. 1972, Shepard et al. 1974, Kogan et al.

1977, Rudd and Jensen 1977, Pitre et al. 1987).

Soybean Looper

Soybean looper is a lepidopteran in the family Noctuidae and subfamily Plusinae. The distribution of this species tends to be limited to the western hemisphere. In the United States, soybean looper ranges throughout most of the country from New York to California, but rarely reaches levels of economic importance outside of the Southeast (Herzog 1980). Soybean looper is a migratory pest and has a limited overwintering region in the United States, primarily confined to the most southern regions of Florida and Texas. Overwintering populations begin to increase in the spring months, as temperatures become more favorable for development, a 6

percentage of moths will migrate northward (Mitchell et al. 1975). Outside of the United States, soybean looper reproduces year round in regions of Central and South America and the

Caribbean where it will subsequently migrate northward as the weather becomes more favorable

(Boethel et al. 1992). However, it is not known if some northern populations migrate back south to contribute to the overwintering populations, or if these wintering populations are exclusively southern populations that expand their range each year (Mason et al. 1989). The soybean looper is a polyphagous species with hosts in more than 28 plant families, but prefers soybean for feeding and oviposition (Moonga and Davis 2016).

Soybean looper adults are relatively small moths with a forewing length of 13 to 18 mm, a robust body with small tufts of scales on the dorsal thorax and anterior abdomen. The forewing is gray-brown with copper iridescence and is darkest just above the stigma and palest from the base to the antemedial line (Eichlin and Cunningham 1978, Lafontaine and Poole 1991). The stigma is oblique and U-shaped accompanied by a silver spot that is roughly the same size and mostly separated from the stigma (Eichlin 1975). The hind wing is dark brown with a white fringe intersected with brown spots (Pogue 2005). Adults typically feed on nectar from flowering crops, the nectar of cotton appears to be important as a source of sugars and carbohydrates which provide the nutrients for egg production and later migration (Jensen et al.

1974). Female adults prefer to oviposit in reproductive stage soybeans, where they lay significantly more eggs than in vegetative stages soybeans or in cotton (Mason et al. 1989,

Felland et al. 1992, Mascarenhas and Pitre 1997, Jost and Pitre 2002). Oviposition takes place when the moths are most active, between dusk and dawn, and they prefer to oviposit in the middle to upper plant canopy (Mascarenhas and Pitre 1997). More eggs tend to be laid on the bottom surface of leaves than the top surface and other plant structures. This may be due to the

increased trichome density found on the lower leaf surface (Jost and Pitre 2002, Mason et al.

1989, Mascarenhas and Pitre 1997). Oviposition peaks at five days after adult emergence and remains high until the 13th day after adult emergence. Approximately 75% of the eggs are laid singly with ca. 72% viability (Mason et al. 1989, Pansera-De-Araujo et al. 1999b). Early planted soybeans, which have greater canopy development than later planted soybeans, and soybeans grown on narrow row spacings are often preferred (Mascarenhas and Pitre 1997). Eggs are globular, approximately 0.02 to 0.5 mm in diameter and are pale yellow with flat and smooth base (Barrionuevo and San Blas 2016). Eclosion of the egg takes three to five days. During this time the egg goes through eight stages of embryonic development: meiosis and fertilization, cytoplasmic retraction, early stages of gastrulation, the 1st stage of germ band development, the

2nd stage of germ bad development, segmentation of the egg, development of several body structures, and the final events of organogenesis. After development, eclosion occurs where nearly half of the larvae eat the chorion (Pansea-De-Araujo et al. 1999a). The soybean looper develops through five to six instars, which is dependent upon the diet makeup early in the life cycle (first through third instar). With 76% of individuals that pupate after the fifth instar while

24% require a sixth when feeding on soybean (Strand 1990). The first instar develops in three to four days, during which it feeds for approximately half of that time and then wanders and molts.

A shed black head capsule is an indicator of the completion of this stage. The second through fourth instars develop in two to three days each, and are all divided into a wandering and feeding stage (Shour and Sparks 1981). During the second and third instar, the larvae undergo a noticeable color change. During feeding, larvae will appear green-brown, while during wandering they will appear translucent lime green. The transition from feeding to wandering is marked by the head being disproportionately smaller than the body. Following gut purge, the

larvae will spin a fine silk mat on the underside of leaves to assist their molt to the next instar

(Shour and Sparks 1981). The fourth instar is more variable in color, from bright green to yellow-green. During this instar, white lines of variable sizes become conspicuous dorsally, addorsally, subdorsally, and laterally (Barrionuevo and San Blas 2016). The penultimate instar is divided into four phases: new, feeding, early head capsule slippage, and late head capsule slippage. The final instar can be divided into five phases: new, feeding, wandering, cocoon- spinning, and pharate pupa. Once the larval stages are complete, the pupae will be encapsulated in a delicate cocoon and pupa are bright green in the first days then will begin to turn light brown as they approach emergence which takes one to two hours (Eichlin 1975, Barrionuevo and San

Blas 2016). Soybean looper larvae consume large, irregular areas of soybean leaves, leaving the large lateral leaf veins intact, which gives the severely defoliated leaves a lace-like appearance

(Herzog 1980). Although all instars of soybean looper larvae feed on soybean, consumption from third through final stage larvae accounts for approximately 95% of total foliage consumption

(Sullivan and Boethel 1994, Trichilo and Mack 1989). Larval feeding typically occurs in the lower 1/3 to 2/3 of the canopy and as populations increase, larvae migrate and consume leaves in the upper canopy (Herzog 1980). Leaf age appears to have little or no effect on feeding, but small larvae tend to feed more selectively on younger leaves, whereas mature larvae are more indiscriminate (Jensen 1974).

Soybean looper can be controlled by a variety of naturally occurring predators throughout their life cycle. As eggs, soybean looper can be parasitized by Trichogramma spp., a minute polyphagous wasps; and Copidosoma truncatellum (Dalman) an egg or larval parasitoid that develops with the host and emerges during the pre-pupal stage (Harding 1976, Daigle et al.

1990). Early instar larvae (first – third) can be parasitized by four species: Mesochorus

discitergis (Say) (Family: Ichneumonidae), Copidosoma truncatellum (Dalman) (Family:

Encyritidae), Cotesia marginiventris (Cresson) and Meteorus autographae (Muesebeck)

(Family: Braconidae) all of which are parasitic wasps. Late instar larvae (third - fifth) can be parasitized by four species: Chaetophlepsis plathypenae (Sabrosky) and Lespesia aletiae (Riley)

(Family: Tachinidae) which are true flies, Euplectrus comstockii (Howard) (Family: Eulophidae), and Cotesia spp.. Pupal stage soybean loopers can be parasitized by Pediobious facialis (Giraud)

(Family Eulophidae) and Brachymeria ovata (Say) (Family: Chalcididae) both parasitic wasps.

In addition to these parasitoids, soybean looper can be infected by various pathogens: a baculovirus and a nuclear polyhedrosis virus (NPV) are the predominant pathogens.

Entomophthora sp. and Nomuraea rileyi (Samson), both pathogenic fungi, also occur (Daigle et al. 1990).

Soybean looper has developed resistance to many classes of insecticide used for its control, including cyclodienes, organophosphates, carbamates, and pyrethroids (Boethel et al.

1992). Economically damaging populations of soybean looper occur more commonly in soybean than in other crops in the southeastern United States. However, selection for resistance seems to be more important in crops other than soybean because soybean is rarely treated more than once per season for the control of the soybean looper (Thomas and Boethel 1994). Selection in overwintering areas is also important. Newsom et al. (1975) proposed that methomyl resistance in soybean loopers collected from soybean in Georgia and South Carolina was a result of selection on vegetables and ornamentals in Florida (Thomas and Boethel 1994). Permethrin was highly effective for soybean looper control when first registered (Boethel et al. 1992), but efficacy has declined such that it is often unsatisfactory (Thomas and Boethel 1993). Permethrin resistance is more severe in cotton-soybean ecosystems, probably because of inadvertent

selection of soybean loopers that results from heavy treatment of cotton with pyrethroids

(Felland et al. 1990, Leonard et al. 1990). In addition to permethrin resistance, soybean looper resistance to most chemicals was higher in strains collected where cotton was more widely grown (Thomas and Boethel 1994, Felland et al. 1990, Leonard et al. 1990) Little is known about the physiological mechanism of insecticide resistance in the soybean looper. Rose et al. (1990) found elevated levels of several metabolic enzymes, while evidence of target site resistance was found by Thomas and Boethel (1994).

Soybean Thresholds

Integrated pest management is based on the premise that certain levels of pests are tolerable (Peterson and Hunt 2003). In integrated pest management the economic injury level

(EIL) and the economic threshold (ET) are the most commonly used decision tools (Pedigo and

Higley 1996). The EIL is a cost-benefit equation where the costs (losses associated with managing a pest) are balanced with the benefits (losses prevented by managing a pest). The most commonly used equation to determine the EIL is: EIL = C / VIDK where C is the management costs per production unit ($ / ha), V is the market value per production unit ($ / ha), I is the injury unit per pest equivalent, D is the damage per unit injury (Kg reduction / ha / injury unit), and K is the proportional reduction in injury with management (Peterson and Hunt 2003). Stern et al. (1959) defined the EIL as “the lowest population density of a pest that will cause economic damage (the amount of injury that justifies the cost of artificial control measures)”. The ET was then defined as “the density at which control measures should be initiated to prevent an increasing pest population from reaching economic injury level” (Stern et al. 1959). Because a time delay often occurs between the time of sampling and the application of a control measure, the ET is set lower than the EIL so that increasing populations cannot exceed the EIL during this 11

delay. Additionally, setting the ET lower than the EIL minimizes the risk of the population exceeding the EIL due to sampling procedure variation, the increase of a zpest population, time delays between sampling and control applications, etc. (Poston et al. 1983).

Throughout the United States, defoliation and soybean looper ET’s vary by state.

Defoliation thresholds tend to be set prior to bloom and after bloom for soybeans, this accounts for the plants ability to compensate for photosynthetic losses prior to reproductive stages.

Defoliation economic thresholds are set based on each state’s economic injury level which varies based on yield potential and value of crop. Additionally, there is no standardized method to determine the ET. In most cases, they tend to be 15 to 25% lower than the EIL.

Defoliation during scouting is usually determined by visual inspection and therefore varies by individual. This often leads to an overestimation of defoliation, particularly with regards to soybean looper which feed on leaf tissue primarily located in the upper 2/3 of the plant. This has led to a number of states establishing population thresholds based on soybean looper’s potential for causing defoliation (Table 1.1).

Soybean loopers are a late-season defoliating pest whose impact on yield largely depends on the soybean’s growth stage at the time of infestation and the density of the infesting population. Throughout the mid-southern United States, thresholds and sampling methods are not standardized for determining defoliation or soybean looper larval densities. The defoliation threshold for reproductive stage soybean is 20% in Mississippi (Owen et al. 2013). This combined with the common practice of using earlier maturing and higher yielding soybean varieties, called the current soybean looper larval threshold into question. Therefore, three objectives were determined to be used in the refinement of the population density threshold. The first objective was to develop an equivalency between the drop cloth and sweep net sampling

methods for soybean looper larvae infesting soybean in Mississippi. The second objective was to evaluate soybean looper larvae consumption rates. The final objective was to further refine and/or validate the soybean looper larval economic threshold for soybean in Mississippi.

Table 1.1 Defoliation and sampling thresholds for soybean looper larvae in soybean producing states.

Defoliation Drop Cloth Threshold Sweep Net Threshold Reference State Threshold (per 1.52 row meters) (per 25 sweeps)

Prior to After Prior to After Prior to After

Bloom Bloom Bloom Bloom Bloom Bloom Alabama 35% 20% 25 – 40b 38b (Reed et al. 2018) Arkansas 40% 25% 30 – 40a 29a (Studebaker et al. 2019) Florida 30% 15% - - (Carter and Gillett-Kaufman 2017) Georgia 30% 15% 40a 19a (Roberts 2019) Kentucky 35% 25% - - (Johnson 2016) Louisiana 35% 25% 40a 38a (Ring et al. 2019) Mississippi 35% 20% 40a 20a 38a 19a (Catchot et al. 2019) North Carolina 30% 15% - - (Reisig 2018) Oklahoma 35% 15% - - (Royer 2019) South Carolina 30% 15% 40a 38a (Greene 2018)

Tennessee 30% 20% - 19a (Stewart and McClure 2019) Texas 40% 20% - - (Vyavhare et al. 2015) Virginia 40% 15% - - (Taylor and Herbert 2019) a 1.27 cm (0.5 in.) or larger b 0.635 cm (0.25 in.) or larger

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CHAPTER II

EVALUATION OF DROP CLOTH AND SWEEPNET SAMPLING METHODS FOR

SOYBEAN LOOPER INFESTING SOYBEAN IN MISSISSIPPI

Abstract

Field experiments were conducted in 2017 and 2018 in multiple commercially grown soybean fields to determine the relationship between the drop cloth and sweep net sampling methods for soybean looper larvae. These studies were conducted across a variety of insect population densities, planting dates, row widths, and plant heights. Multiple samples per location were collected using each method. Multiple samplers were utilized to get an estimate of sampler variability. Overall, the sweep net method was more effective at capturing soybean looper larvae regardless of size than the drop cloth sampling method. The sweep net method was more effective at capturing larvae third instar or greater when compared to larvae less than third instar.

Introduction

In soybean, the drop cloth technique for sampling is widely recommended for sampling defoliators and pod feeders (Strayer and Greene 1974, Barnes et al. 1974, Kogan et al. 1977).

Despite this recommendation, the sweep net remains a widely used tool for sampling arthropods on many row crops, including soybean. Measurements taken to estimate populations in field crops fall into three groups: absolute methods, relative methods, and population indices.

Absolute methods provide estimates of density per unit of land area. Relative methods give density per some unit other than land area. Population indices do not count insects at all, but 22

rather measures effects of insect feeding on plants, such as defoliation (Ruesink 1980). The drop cloth provides absolute population estimates for lepidopteran species and the sweep net is a relative sampling method (Kogan and Pitre 1980). For most pest management purposes determining relative population estimates that permit a rapid classification into decision categories (e.g., spray or do not spray) is usually sufficient (Ruesink and Kogan 1975). The procedure for sampling with the drop cloth consists of stretching the cloth between two adjacent rows of plants so that the edges of the cloth touch the plants. The surveyor then bends the plants of both adjacent rows within the length of the drop cloth over the cloth and beats the plants to dislodge arthropods. Narrow row spacing limits the ability to completely unravel the drop cloth and bend the plants over the cloth without disturbing the adjacent row. Even when used by different surveyors, the drop cloth produces reasonably consistent results because the main factor affecting the procedure is the vigor of the shaking action. A weaker shake can be compensated for by increasing the number of shakes (Kogan and Pitre 1980).

Despite difficulties in standardizing the sampling procedure, the sweep net has been the most widely used tool for sampling arthropods on small grain, forage, and row crops for over a century (Kogan and Pitre 1980). This popularity is due to the fact that no other method can capture as many insects from vegetation per man hour without increased cost for equipment and damage to the crop (Ruesink and Kogan 1975). A sweep net user walks alongside a row, swinging the net through the foliage in a figure-8 pattern as they progress, counting each pass through the canopy as one sweep (Kogan and Pitre 1980). There are a number of environmental factors that can contribute to variability in sampling numbers including temperature, humidity, wind velocity, position of the sun, plant size, density of the canopy, pubescence of leaves, height of sweeping, and sweeping method. Each of these factors can affect either the surveyor’s ability

to effectively sweep the canopy or the insect’s escape reaction (Delong 1932, Kogan and Pitre

1980). Previous research by Rudd and Jensen (1977) paired 1.82 row meter drop cloth samples with 25 sweeps taken with a 38 cm sweep net in soybeans that were in full-bloom until harvest.

Results were paired as the number caught in a single drop cloth measurement and the number caught per 25 sweeps. Additionally, the mean of number caught per drop cloth per field were paired with the mean of the numbers caught per sweep net sample per field. When comparing single drop cloth samples with single sweep net samples results do not show a strong correlation which could be due to within-field variability of insect populations. When comparing field means of drop cloth samples with field means of sweep net samples the correlation is very good.

For both, the single observations and field mean observations results show that estimates obtained by one method are nearly equal to estimates obtained by the other method (Rudd and

Jensen 1977). Marston (1979) converted the number of larvae collected by drop cloth samples to whole plant sampling. On 76 cm rows with an open canopy, a 30 cm section of row was sampled with a drop cloth. In that study, a predetermined number of larvae were placed in those sections prior to sampling. Because the number sampled and the number present were known, a recovery percentage could be calculated. In this study, 76.5% of small larvae (first and second instar),

95.9% of medium larvae (third and fourth instar), and 97.7% of large larvae (fifth and sixth instar) were captured. Marston (1979) reported that a greater percentage of larvae were recovered as plants increased in growth stage, when sampling a population of known size. Studebaker et al.

(1991) conducted a field cage study comparing population estimates by drop cloth and sweep net samples at three soybean growth stages (R2, R4, and R5-6) which were planted on either 84 or

96 cm rows. In some cages, four 1.82 row meter drop cloth samples were collected and expressed as numbers of larvae per 0.81 row meters. Sweep net samples were collected in cages

with a 38 cm sweep net, 3 previously unsampled rows in each cage were sampled with six sweeps per row and were expressed as number of larvae per 0.38 row meters (1 sweep). Neither method gave an absolute estimate of populations levels. At the R2 growth stage, sampling with the drop cloth was more effective at collecting third, fourth, and sixth instar soybean looper larvae, sweep net sampling was more effective at collecting fifth instar larvae, but only slightly.

At the R4 growth stage, fourth, fifth, and sixth instar larvae were all more effectively sampled with the drop cloth than with the sweep net. At growth stage R5-6, third though sixth instar larvae were all more effectively sampled with the drop cloth than with the sweep net.

The establishment of reliable conversions from one method of sampling to another is integral to the creation of an integrated pest management plan. In research capacities, conversions can be used to convert counts from caged plant studies (which are often too small to sample with anything other than a drop cloth) to counts that are compatible with threshold numbers. Therefore, a study was designed to obtain conversions between the sweep net and drop cloth sampling methods.

Materials and Methods

Experiments were conducted in 2017 and 2018 in commercial soybean fields throughout

Mississippi and at the Delta Research and Extension Center in Stoneville, MS. In 2017, commercial soybean fields in Flora, MS were sampled on 3 August by eight individuals; in

Canton, MS on 14 August by two individuals; in Stoneville, MS on 25 August by four individuals; and in Belzoni, MS on 16 August by two individuals. In 2018, samples were collected in Epps, LA on 3 August by ten individuals; in Glendora, MS on 9 August by nine individuals, and in Webb, MS on 16 August by four individuals. In Stoneville, MS, two locations were sampled for this experiment. The first location was sampled on 4 September and again 14 25

September, both times by four individuals. The second location was sampled on 4 September, 14

September, and 19 September, each time by four individuals. Environmental conditions that could cause variation in samples were recorded at the time of sampling using the nearest weather station (NOAA 2019) (Table 2.1). During both years, samples were collected in a similar manner. Individuals sampled various locations throughout naturally infested fields by using 38.1- cm diameter sweep net and a 0.76-m long drop cloth. Each individual sampled a 4 to 6 row section along one side of each field, careful to sample only previously unsampled rows. After completing 5 sets of 25 sweeps, 5 drop cloth samples were taken no more than three rows from where the sweep-net samples were collected, similar to Deighan et al. (1985). Drop cloth samples were obtained by carefully unrolling a 0.76m drop cloth between two adjacent rows until the edges of the drop cloth touched the bases of the plants. From there, plants on both sides of the drop cloth were bent over the drop cloth and vigorously shaken to dislodge as many larvae as possible. Soybean looper larvae were counted and classified by size for each sampling method. In 2017, larvae were classified as either less than 1.27 cm (0.5 in, less than third instar) or greater than or equal to 1.27 cm (0.5 in, greater than or equal to third instar). In 2018, larvae were classified as either: less than third instar (approx. less than 1.27 cm) or as third, fourth, or fifth instar.

At each location, samples were averaged by individual to account for any variation in the sampling procedure. The relationship between the number of soybean looper larvae captured using sweep net and drop cloth sampling was determined with regression analysis using the SAS

Glimmix procedure (Version 9.4, SAS Institute, Cary, NC). The relationship between the number of larvae captured using each method was determined for less than third instar larvae, third, fourth, and fifth instar larvae, ≥ third instar larvae, and total larvae. Siteyear (combination

of year and location) and sampler nested in siteyear were treated as random effects. The variable siteyear refers to each location by date where samples were taken with sampler acting as replications. Degrees of freedom were calculated using the Kenward-Roger method. R-squared was calculated using the SAS Reg procedure (Version 9.4, SAS Institute, Cary, NC).

Results

Significant relationships for numbers of less than third instars (F = 4.60 , df =1,22 , P =

0.0432 , R2 = 0.1053 ), third instar (F = 11.30 , df = 1,52.2 , P = 0.0015 , R2 = 0.2464 ), fourth instar (F = 75.21 , df = 1,37.18 , P < 0.0001 , R2 = 0.6921 ), fifth instar (F = 110.42 , df =

1,24.76 , P < 0.0001 , R2 = 0.6360 ), greater than or equal to third instar (F = 33.07 , df = 1,46.93

, P < 0.0001 , R2 = 0.5868 ), and total soybean looper larvae (F = 3.40, df = 1,52.91 , P = 0.0707,

R2 = 0.0745 ) captured using sweep net and drop cloth sampling were observed. Intercept term was significant for less than third, third, fourth, greater than or equal to third instar, and total instar. The intercept term for fifth instar was not significant (P = 0.0548), therefore the no intercept (NOINT) option was used for the fifth instar relationship.

Based on the regression equation, for every soybean looper larvae less than third instar captured with drop cloth sampling, 3.49 were captured using sweep net sampling (Figure 2.1).

While for every third instar larvae captured using drop cloth sampling, 2.74 were captured using a sweep net (Figure 2.2). For fourth instar larvae, every larvae captured using the drop cloth sampling method equates to 2.02 larvae captured using the sweep net sampling method (Figure

2.3). For every fifth instar larvae captured using the drop cloth, 1.30 larvae were captured using the sweep net sampling method (Figure 2.4). Every larvae greater than or equal to third instar captured using the drop cloth, 4.77 larvae were captured using the sweep net sampling method

(Figure 2.5). For total soybean looper larvae, for every larvae captured using the drop cloth,

11.52 larvae are captured using the sweep net method.

Discussion

Several studies have investigated the relationship between drop cloth sampling and sweep net sampling either using natural infestations, or by infesting known numbers of larvae in cages and calculating the recovery percentage for the sampling method tested (Shepard et al. 1974,

Marston et al. 1979, Studebaker et al. 1991). With natural infestations of soybean looper and cabbage looper, Trichoplusia ni (Hübner), the drop cloth method was generally captured more less than third and third instar or greater soybean looper larvae than the sweep net method.

(Shepard et al. 1974) With known densities of soybean looper larvae, larval recovery using drop cloth sampling varied with larval size and soybean growth stage.

As larvae increased in size, the recovery percentage increased. Recovery percentage also increased as growth stage increased (Marston et al. 1979). Using field cages and known larval densities, larval recovery with drop cloth sampling generally increased as soybean plants progressed through the reproductive growth stages, while larval recovery using sweep net sampling did not (Studebaker et al. 1991). Recovery percentages for sweep net sampling were generally lower than for drop cloth sampling. One study directly compared the two sampling methods. For total soybean looper larvae, drop cloth sampling was more effective than sweep net sampling, and captured 12.4 soybean looper larvae (per 1.82 row m) for every 10 (per 25 sweeps) captured using sweep net sampling (Rudd and Jensen 1977). In the current study, drop cloth sampling was more effective in capturing less than third and third instar soybean looper larvae compared to sweep net sampling. While sweep net sampling was more efficient than drop cloth sampling in capturing fourth, fifth, total larvae greater than or equal to third instar, and total 28

soybean looper larvae. One major difference between the current study and the previous studies mentioned above is the type of soybean variety used in the experiment. Determinate maturity group VI and VII soybean varieties were used in the previous studies and reflected the varieties being grown by producers during that time. In the period between the previous studies and the current study, growers in the mid-southern U.S. have transitioned to indeterminate maturity group IV and V soybean varieties that are generally shorter in stature than MG VI and VII varieties. These differences in growth habitats, particularly plant height, may explain the differences in larval capture efficiencies between the studies. In many states, the soybean looper larval threshold considers only larvae greater than or equal to third instar. In the current study capturing 10 soybean looper larvae greater than or equal to third instar per 1.52 row meters using drop cloth sampling was equivalent to capturing 12.57 larvae per 25 sweeps. This information is useful to growers or consultants who prefer drop cloth sampling and no threshold number for drop cloth sampling is listed. Also, these data would have utility for researchers using field cages. Plants within cages could be sampled using a drop cloth and converted to an equivalent sweep net capture.

The defoliation threshold for reproductive stage soybeans in Mississippi is 20% (Owen et al. 2013). With the recent establishment of this threshold, it is unclear how the current sampling methods for soybean looper larvae relate to the defoliation threshold. As part of the process for determining whether the current sampling threshold accurately predicts the number of larvae that will cause 20% defoliation damage we first wanted to establish how each current sampling method threshold related to each other. Currently for reproductive stage soybeans in Mississippi.

The threshold for soybean looper larvae sampled with a drop cloth is 20 larvae greater than or equal to 1.27 cm (0.5 in) per 1.524 row meters. Using the regression equation Y = 0.2141x +

11.302 (for the total number of larvae, regardless of size) the current drop cloth sampling threshold equates to 16 larvae per 25 sweeps which is slightly lower than the 19 larvae per 25 sweep threshold that is published (Catchot et al. 2019)

Table 2.1 Environmental data, row spacing, and crop height for sampling sites during 2017 and 2018. Row Spacing Crop Height Date Location Temperature (°C) Humidity (%) Sampling Time (cm) (cm)

3 Aug 2017 Flora, MS 30 84 14:00 76 91

14 Aug 2017 Canton, MS 32 61 12:30 97 102

16 Aug 2017 Belzoni, MS 33 78 13:30 97a 91

25 Aug 2017 Stoneville, MS 28 78 11:30 102 89

3 Aug 2018 Epps, LA 36 68 10:30 97 97

9 Aug 2018 Glendora, MS 32 24 10:00 97 94

16 Aug 2018 Webb, MS 34 60 10:30 97a 91

4 Sep 2018 Stoneville, MS (A) 32 61 13:30 102 109

4 Sep 2018 Stoneville, MS (B) 32 59 14:45 102 114

14 Sep 2018 Stoneville, MS (A) 33 59 14:15 102 109

14 Sep 2018 Stoneville, MS (B) 33 58 15:30 102 114

19 Sep 2018 Stoneville, MS (B) 36 40 14:30 102 114 a twin row planted soybean

Figure 2.1 Relationship between numbers of soybean looper larvae less than third instar captured using drop cloth and sweep net sampling.

Figure 2.2 Relationship between numbers of third instar soybean looper larvae captured using sweep net and drop cloth sampling.

Figure 2.3 Relationship between numbers of fourth instar soybean looper larvae captured using drop cloth and sweep net sampling.

Figure 2.4 Relationship between numbers of fifth instar soybean looper larvae captured using sweep net and drop cloth sampling.

Figure 2.5 Relationship between numbers of soybean looper larvae greater than or equal to third instar captured using drop cloth and sweep net sampling.

Figure 2.6 Relationship between total number of soybean looper larvae captured using drop cloth and sweep net sampling methods.

References Cited

Barnes, G., B. F. Jones, and W. P. Boyer. 1974. Control insects on soybeans. 6pp. Leafl. 193. Arkansas Agric. Ext. Serv. University of Arkansas. Fayetteville, AR.

Deighan, J., R. M. McPherson, and F. W. Ravlin. 1985. Comparison of sweep-net and ground-cloth sampling methods for estimating arthropod densities in different soybean cropping systems. J. Econ. Entomol. 78: 208 – 212.

DeLong, D. M. 1932. Some problems encountered in the estimation of insect populations by the sweeping method. Ann. Entomol. Soc. Amer. 25: 13-17.

Kogan, M., S. G. Turnipseed, M. Shepard, E. B. de Oliveira, and A. Borgo. 1977. Pilot insect pest management program for soybean in Southern Brazil. J. Econ. Entomol. 70: 659-663

Kogan, M., and H. N. Pitre, Jr. 1980. General sampling methods for above-ground populations of soybean arthropods, pp 30 – 60. In M. Kogan and D. C. Herzog (ed.), Sampling Methods in Soybean Entomology, 1st ed. Springer-Verlag New York Inc.

Marston, N. L., W. A. Dickerson, W. W. Ponder, and G. D. Booth. 1979. Calibration ratios for sampling soybean lepidoptera: effect of larval species, larval size, plant growth stage, and individual sampler. J. Econ. Entomol. 72: 110-114.

National Oceanic and Atmospheric Administration (NOAA). 2019. Weather and hazards data viewer, historical surface observations. National Weather Service. May 10, 2019.

Owen, L. N., A. L. Catchot, F. R. Musser, J. Gore, D. Cook., R. Jackson, and C. Allen. 2013. Impact of defoliation on yield of group IV soybean in Mississippi. Crop Prot. 54: 206-212

Rudd, W. G., and R. L. Jensen. 1977. Sweep net and ground cloth sampling for insects in soybeans. J. Econ. Entomol. 70: 301-304.

Ruesink, W. G. 1980. Introduction to sampling theory, pp 61 – 78. In M. Kogan and D. C. Herzog (ed.), Sampling Methods in Soybean Entomology, 1st ed. Springer-Verlag New York Inc.

Ruesink, W. G. and M. Kogan. 1975. The quantitative basis of pest management: sampling and measuring, pp 355-392. In R. L. Metcalf and W. H. Luckman (ed.), Introduction to Insect Pest Management. John Wiley and Sons New York.

Shepard, M., G. R. Carner, and S. G. Turnipseed. 1974. Seasonal abundance of predaceous arthropods in soybeans. Environ. Entomol. 3: 985 – 988.

Strayer, J. and G. L. Greene. 1974. Soybean insect management, 9 pp. Circ. 395. Florida Coop. Ext. Serv. University of Florida. Gainesville, FL.

CHAPTER III

EVALUATION OF SOYBEAN LOOPER LARVAL CONSUMPTION RATES

Abstract

Laboratory experiments were conducted in 2018 and 2019 to determine the feeding rates of soybean looper larval instars on MG IV soybeans. Data collected in this study were obtained using greenhouse grown leaves and a lab colony of soybean looper larvae at least 50 generations removed from the field. Leaf weight, area, and larval weight were recorded before and after feeding occurred. Leaf area consumption by less than third instar larvae was less than feeding from larvae greater than or equal to third instar, with fifth instar larvae consuming the most leaf area. Leaf consumption by weight for larvae less than or equal to third instar was similar to consumption by fourth instars, with fifth instar consuming the most leaf tissue by weight. The largest amount of weight gain occurred during the fourth instar, with the next highest amounts occurring during the third and fifth instar. Data from Reid and Greene (1973), Kogan and Cope

(1974), Boldt et al. (1975), and Trichillo and Mack (1989) were used in combination with data recorded in this study to create a range of leaf consumption for each larval instar. From these ranges, three feeding models (aggressive, moderate, and conservative) were created to represent various scenarios in which soybean looper larval feeding could occur. These models and their respective insect equivalent calculations could then be used in further analysis to refine the soybean looper larval sampling threshold.

Introduction

Soybean looper, Chrysodeixis includens (Walker), larval feeding rate for different developmental stages (instars) are important for developing accurate thresholds and monitoring insect populations. Feeding rates can be used to determine which developmental stages cause significant amounts of damage, the damage potential of a pest,etc. Soybean looper larvae in

Mississippi usually begin feeding on foliage about the time that soybean reaches full bloom (R2); late August – early September (Fehr et al. 1971).

One study was conducted using field grow leaflets of blooming soybean and soybean looper larvae from a laboratory colony. Larvae completed six instars during their development.

Each larva consumed an average of 81.96 cm2 of leaf area from the first instar through pupation.

First through six instars consumed an average of 0.25, 0.83, 1.77, 7.93, 15.50, and 55.75 cm2, respectively. Consumption during the first three instars comprised only 3.3% of the total consumption, the remaining 96.7% of consumption was completed by the final three instars

(Reid and Greene 1973). Boldt et al. (1975) completed a similar study, but with mature soybean leaves. They found that total mean consumption for one larva was 113.82 cm2 as it matured through six instars. Larvae consumed an average of 0.13, 0.41, 1.99, 5.51, 12.18, and 93.48 cm2 for instars one through six respectively. Sixth instars consumed 82% of total leaf area and fifth instars consumed 10.6%. Total leaf area consumption per larvae reported by Boldt et al. (1975) was 37% greater than the total leaf area consumption per larvae reported by Reid and Greene

(1973). Trichillo and Mack (1989) conducted a feeding study using leaves from pre-flowering soybean plants grown in a greenhouse and a laboratory colony of soybean looper larvae that matured through six instars. Larvae were kept at four temperatures for the duration of their feeding (15.6°C, 21.1°C, 26.7°C, and 32.2°C). As temperature increased, feeding increased up to

26.7°C were feeding peaked, before decreasing at 32.2°C. Kogan and Cope (1974) reported a total of 206.71 cm2 of leaf tissue consumed for their study conducted using a laboratory colony of soybean looper larvae and greenhouse grown leaves. First through sixth instar larvae consumed an average of 1.18, 3.36, 13.81, 26.35, 76.44, and 85.66 cm2, respectively. In this study, 91.1% of leaf area was consumed in the last three instars. As previous studies were conducted using various methods of measuring leaf area as well as older varieties of soybean, a study was conducted to evaluate the amount of leaf tissue consumed by weight and by area per instar, as well as the amount of weight gained per instar using currently available soybean cultivars.

Materials and Methods

In 2017 and 2018, attempts were made to obtain field collections of soybean looper larvae. In 2017, collections were made at 4 locations and in 2018, collections were made at 5 locations (Table 3.1). Individuals were collected with a sweep net in commercial soybean fields.

Larvae greater than or equal to third instar were kept and placed in a rearing room which was kept between 26.7 and 29.4°C and 70% humidity, then monitored for pupation. Collections experienced a high level of mortality or did not produce viable eggs (Table 3.1). The inability to maintain a colony of soybean looper larvae resulted in this study being conducted using larvae obtained from Benzon Research (Benzon Research Inc., PA). This colony was over 50 generations removed from the field and completed their development in five instars. The leaves in this feeding study were obtained from greenhouse grown plants (Asgrow 46X6, Bayer Crop

Science, St. Louis, MO).

An experiment was conducted during 2018 and 2019 to determine the feeding rates of the individual instars of soybean looper larvae. Fifty 10 cm petri dishes were prepared with a piece 42

of filter paper that was dampened with 1 ml of water. Fifty young soybean leaves were then weighed using a standard analytical balance (Mettler-Toledo®, Columbus, OH) and leaf area was measured with a Li-Cor 3000 leaf area meter (Li-Cor Biosciences®, Lincoln, NE). Leaves were measured three times and an average was taken of the three measurements to reduce variation from the leaf area meter. One leaf was placed in each petri dish and half of these petri dishes were infested with one neonate soybean looper larvae which was allowed to feed until molting and the other half of the dishes were left uninfested as a control. Following feeding all leaves were weighted and leaf area was measured again. This process was repeated again for each subsequent instar, with the addition of larval weights before and after feeding for larvae second instar or greater. Larval weights were not recorded for the first instar larvae due to the insensitivity of the scales in relation to the weight of the larvae. This process was repeated to achieve three replications. Controls from all instars and replications were combined and averaged. These averages were then used to correct for control to account for water loss. Data were then analyzed using analysis of variance procedures (PROC GLIMMIX of SAS®) (Version

9.4, SAS Institute, Cary, NC). Instar served as the fixed effect in the model and replication was treated as the random effect. Means were calculated using the LSMEANS statement and separated based on Fisher’s least-significant-difference test for leaf area consumption, leaf weight consumption, and weight gain per instar respectively.

Feeding rates were then compiled with previously published data from: Reid and Greene

1973, Kogan and Cope 1974, Boldt et al. 1975, and Trichillo and Mack 1989 to create a range of feeding rates. These feeding rate ranges were then used to create three models of feeding: conservative, moderate, and aggressive. The conservative feeding model was created using the lowest recorded feeding rate from all of the studies, the moderate feeding model was created

using the mean feeding rate from all of the studies, and the aggressive feeding model was created using the highest feeding rate from all of the studies (Table 3.2). After the models were established, insect equivalents could be calculated, insect equivalents for various instars of the same species are used to compensate for the differences in leaf area consumption of various sized larvae.

Insect equivalents were calculated by comparing each instars leaf area consumption to that of the first instar. By assigning the first instar an insect equivalent of one we considered each instar in relation to the first, each subsequent instar then consumes x times more leaf tissue than the first instar. Using this method allows for larger larvae, which consume more leaf tissue than smaller larvae to be given more statistical weight.

Results

Significant differences in leaf area consumption were observed among soybean looper larvae of different developmental stages (F=35.81, df = 4, 230, P < 0.01) (Figure 3.1). First and second instar soybean looper larvae consumed similar amounts of leaf area, 1.72 and 1.19 cm2, respectively. Leaf area consumption by third (4.66 cm2) and fourth instar larvae (3.87 cm2) was similar and significantly greater than that for both the first and second instar larvae. Fifth instar larvae consumed more soybean leaf area (9.52 cm2) than larvae of other age classes.

Larval instar duration was also determined, with the first and fifth instar lasting a duration of 3 days, while a duration of 2 days was observed for the second, third, and fourth instar. Larval duration in this study is similar to results published by Shour and Sparks (1981).

First and second instar soybean looper larvae consumed similar amounts of leaf area per day

(0.57 cm2 and 0.60 cm2, respectively). Third and fourth instar larvae consumed similar amounts of leaf area per day (2.33 cm2 and 1.93 cm2, respectively), but significantly greater than that 44

consumed by less than third instar larvae. Similar to the total leaf area consumed, fifth instar larvae consumed significantly greater leaf area per day (3.1726 cm2), than the per day amount for larvae of other age classes (Figure 3.2).

Significant differences in consumption of leaf mass was observed among larvae of different age and developmental classes (instars) (F= 45.75, df = 4,230, P < 0.01) (Figure 3.3).

Less than third and third instar larvae consumed similar amounts of leaf mass (28.89 mg and

19.66 mg respectively). Third and fourth instar consumed similar amounts of leaf mass (50.44 and 72.6 mg respectively), with fourth instar larvae consuming significantly greater leaf mass than larvae less than third instar. Similar to leaf area consumption, fifth instar larvae consumed significantly greater leaf mass than larvae of other age classes (216.83 mg).

Differences were observed for larval weight gain during the second through fifth instars

(F = 45.75, df = 4,230, P<0.01) (Figure 3.4). Second instar larvae gained the least amount of body weight (4.34 mg). Third instar larvae gained 15.66 mg. Fourth instar larvae gained significantly more weight than larvae of other age classes (26.95 mg). Fifth instar larvae gained

19.53 mg, which was similar to the weight gained for third instar larvae.

The conservative feeding model (Table 3.3) is comprised of feeding rates at the lowest end of the range established in Table 3.2. First, second, and third instar feeding rates were 0.07,

0.26, and 0.65 cm2, respectively (Trichillo and Mack 1989). First and second instar feeding were combined into a single feeding rate of 0.33 cm2, labeled as less than third. Fourth instar feeding was 3.87cm2 (obtained from this study). Fifth and sixth instar feeding was 6.87 and 8.51 cm2

(Trichillo and Mack 1989). Once the feeding rates were established, insect equivalents could be calculated. Less than third instar were assigned an insect equivalent of one; then by comparing the amount of leaf tissue consumed by each subsequent instar to the amount of leaf tissue

consumed for the less than third instar, insect equivalents could be calculated. For this model of feeding the insect equivalents were 1, 1.97, 11.73, 21.82, and 25.79 for less than third through sixth instar respectively. For example, less than third instar larvae in this model consumed

0.33cm2 of leaf tissue and were assigned an insect equivalent of 1, third instar larvae consumed

0.65 cm2 of leaf tissue, therefore third instar larvae have an insect equivalent of 1.97 because second instar larvae consumed 1.97 times more leaf area than less than third instar larvae.

The moderate feeding model (Table 3.4) obtained by using a mean of the ranges established in Table 3.2 lists feeding rates as 1.88, 4.58, 9.62, 24.10, and 79.67 cm2 for less than third through sixth instar. Insect equivalents for the moderate feeding model obtained by using a mean of the rages is 1, 2.44, 5.12, 12.82, and 42.38 for less than third through sixth instar larvae.

The aggressive feeding model (Table 3.5) obtained by using the highest feeding rate from the ranges established in Table 3.2. Feeding rates for less than third instar were 5.08cm2

(obtained from this study and Kogan and Cope 1974). Third through sixth instar larvae had a feeding rate of 13.81, 26.35, 76.44, and 160.92 cm2, respectively (Kogan and Cope 1974). Insect equivalents for this feeding model were 1, 2.72, 5.19, 15.05, 31.68 for less than third through sixth instars.

Discussion

Multiple studies have been conducted on the topic of soybean looper larvae consumption rates including: Reid and Green 1973, Kogan and Cope 1974, Boldt et al. 1975, and Trichillo and

Mack 1989. In these studies, larvae completed six instars of development, whereas in our study larvae completed only five. To prevent skewing data consumption rates for sixth instar larvae were only obtained from the previously conducted studies. Two studies (Reid and Green 1973,

Boldt et al. 1975) conducted their studies using either leaves grown in the field or mature leaves; 46

leaves from these conditions are often thicker than leaves grown in a greenhouse environment which could lead to variation in feeding rates. All studies were conducted using a laboratory colony of soybean looper larvae, the colony used in our study was 50+ generations removed from the field; it is unknown how many generations removed from the field the colonies were in previous studies. Colonies removed from the field for numerous generations could be subject to feeding patterns that are variable to patterns observed in field. In many situations colonies exclusively feed on artificial diet for many generations and will not transition smoothly into feeding on vegetation.

Results obtained from our study were compared with results obtained from published literature. For first instar larvae, the feeding rate obtained from our study was 1.72 ± 0.19 cm2 which was 0.54 cm2 more than the next highest published result of 1.18 cm2 (Kogan and Cope

1974). Results for second instar larvae indicate that our feeding rate of 1.19 ± 0.15 cm2 was 2.17 cm2 lower than the feeding rate of 3.36 cm2 published by Kogan and Cope (1974) and 0.36 cm2 higher than the feeding rate published by Reid and Greene (1973). The third instar feeding rate of 4.66 ± 1.37 cm2 in the current study was 9.15 cm2 lower than results published by Kogan and

Cope (1974) and 2.67 cm2 higher than results published by Boldt et al. (1975). For fourth instar larvae, our results of 3.87 ± 0.21 cm2 were 0.56 cm2 less than results published by Trichillo and

Mack (1989). Fifth instar larvae feeding rate of 9.52 ± 0.90 cm2 was 2.65 cm2 greater than results published by Trichillo and Mack (1989) and 2.66 cm2 less than results published by Boldt et al.

(1975). Overall, for larvae greater than third instar, our results tended to be the lowest or next to lowest feeding rate when compared with published data. As our colony did not mature through the sixth instar feeding rates were only obtained from previously published data.

Variations in feeding rates can be attributed to a variety of factors including, but not limited to: soybean looper larvae colony variation, variations in environmental conditions, variation in leaf thickness or trichome densities, or variability in leaf area measurement techniques. In the current study, there were multiple attempts to establish field collections.

However, poor survival and low reproduction rates of these collections results in the use of a lab colony. Had it been possible t ouse field collections in the current study, measured feeding rates may have been higher.

Various feeding models were constructed from the compilation of feeding rates.

Conservative, moderate, and aggressive feeding models were created to account for a variety of situations that may be encountered when determining the amount of soybean looper larvae would be necessary to reach an economic injury level. Using an aggressive feeding model would result in a fewer number of larvae causing injury of economic importance, while using a conservative model would predict the opposite.

Table 3.1 Date, location, number of individuals collected, and mortality rates for 2017 and 2018 collections

a Larvae survived through pupation, but failed to produce viable eggs

Table 3.2 Soybean looper larval feeding rates from this study compared with results from (Reid and Green 1973, Kogan and Cope 1974, Boldt et al. 1975, Trichillo and Mack 1989).

Consumption rate (cm2) ± SE

Source First Instar Second Instar Third Instar Fourth Instar Fifth Instar Sixth Instar

Trichillo and Mack 1989 0.07 ± 0.01 0.26 ± 0.06 0.65 ± 0.07 4.43 ± 0.92 6.87 ± 0.88 8.51 ± 0.84

Boldt et al. 1975 0.13 ± 0.001 0.41 ± 0.001 1.99 ± 0.11 5.51 ± 0.47 12.18 ± 0.78 93.48 ± 4.34

Kogan and Cope 1974 1.18a 3.36a 13.81a 26.35a 76.44a 160.92a

Reid and Greene 1973 0.25a 0.83a 1.77a 7.93a 15.50a 55.75a

Current Study 1.72 ± 0.19 1.19 ± 0.15 4.66 ± 1.37 3.87 ± 0.21 9.52 ± 0.90 a Standard error not available

Table 3.3 Conservative, moderate, and aggressive feeding rate and insect equivalent models for soybean looper larvae feeding on soybean leaves

Conservative Feeding Model Moderate Feeding Model Aggressive Feeding Model

Instar Consumption Insect Consumption Insect Consumption Insect

(cm2) Equivalent (cm2) Equivalent (cm2) Equivalent

Less than Third 0.33a 1 1.88d 1 5.08bc 1

Third 0.65a 1.97 4.58d 2.44 13.81b 2.72

Fourth 3.87c 11.73 9.62d 5.12 26.35b 5.19

Fifth 6.87a 21.82 24.10d 12.82 76.44b 15.05

Sixth 8.51a 25.79 79.67d 42.38 160.92b 31.68 a Trichillo and Mack 1989 b Kogan and Cope 1974 c Current study d Results are an average from all sources

References Cited

Boldt, P. E., K. D. Biever, and C. M. Ignoffo. 1975. Lepidopteran pests of soybeans: consumption of soybean foliage and pods and development time. J. Econ. Entomol. 68: 480-482.

Fehr, W. R., C. E. Caviness, D. T. Burmood, and J. S. Penington. 1971. Stages of development descriptions for soybeans, Glycine max (L.) Merr. Crop. Sci. 11: 929-931.

Kogan, M. and D. Cope. 1974. Feeding and nutrition of insects associated with soybeans. 3. food intake, utilization, and growth in the soybean looper, Pseudoplusia includens. Ann. of the Entomol. Soc. of Am. 67: 66-72

Reid, J. C. and G. L. Greene. 1973. The soybean looper: pupal weight, development time, and consumption of soybean foliage. Fl. Entomol. 56: 203-206

Shour, M. H. and T. C. Sparks. 1981. Biology of the soybean looper, Pseudoplusia includens: characterization of last-stage larvae. Ann. Entomol. Soc. Am. 74: 531-535

Trichilo, P. J., and T. P. Mack. 1989. Soybean leaf consumption by the soybean looper (Lepidoptera: Noctuidae) as a function of temperature, instar, and larval weight. J. Econ. Entomol. 82: 633-638.

CHAPTER IV

REFINEMENT AND VALIDATION OF SOYBEAN LOOPER LARVAL ECONOMIC

THRESHOLD FOR SOYBEANS IN MISSISSIPPI

Abstract

Soybean looper defoliation was observed for 2017 and 2018 in naturally infested field trials. Data from these field trials were used to show the change in defoliation over ten days.

Feeding models from Chapter 3 were combined with data from this chapter to construct three prediction models for the R3, R5 and R6 economic injury levels published in Owen (2012) and

Owen et al. (2013) Based on these data, the conservative model predicts that for every fifth instar larvae sampled, defoliation would increase 0.75%; the moderate feeding model predicts that for every fifth instar larvae sampled, defoliation would increase 1.27%; the aggressive feeding model predicts that for every fifth instar larvae sampled, defoliation would increase 1.31%.

Using regression equations, economic injury levels (percent defoliation) were converted to insect equivalents, then by using the feeding models presented in Chapter 3, insect equivalents could be converted back into larval samples to equate to the percent defoliation thresholds. Under the current 25% defoliation EIL (the 20% defoliation threshold is 80% of this) at the R3 growth stage this would equate to 58 third instar larvae, 10 fourth instar larvae, or 5 fifth instar larvae under the conservative model; 28 third instar larvae, 13 fourth instar larvae, or 5 fifth instar larvae under the moderate model; and 27 third instar larvae, 14 fourth instar larvae, or 5 fifth instar larvae under the aggressive model. At the R5 growth stage this would equate to 7 third

instar larvae, 11 fourth instar larvae, or 6 fifth instar larvae under the conservative model; 30 third instar larvae, 14 fourth instar larvae, or 6 fifth instar larvae under the moderate model, or 30 third instar larvae, 16 third instar larvae, or 5 fifth instar larvae under the aggressive model.

Introduction

Soybean defoliation can be described in two patterns: (1) a gradual removal of leaf tissue, typically not exceeding 25% of the total leaf area, and occurring before reproductive growth stages; (2) more severe defoliation of 50 to almost 100% by pests that occurs with a 14-day period during reproductive growth stages (Herbert et al. 1992). However, defoliation is not limited to these two patterns. In regions where soybean is produced year round, soybean looper can be an economic pest during both vegetative and reproductive stages. Defoliation may reduce transpiration and photosynthesis in the plant as well as decreasing the plant’s capacity to compensate for nutrient deficiencies, water loss, and other abiotic factors all leading to reductions in soybean yield (Fehr et al. 1985). In pest management situations, percent defoliation is commonly used to predict yield losses based on leaf area removed and initial canopy size

(Klubertanz et al. 1996).

Since 2007, anywhere from 33 to 85% of Mississippi soybeans have been infested with soybean looper, Chrysodeixis includens (Walker), with 14 to 58% of soybeans reaching infestation levels of economic importance (Musser et al. 2008 – 2018). Soybean looper is primarily a defoliating pest that causes damage by consuming large, irregular areas of soybean leaves, leaving the large lateral leaf veins intact which give infested areas a lace-like appearance

(Herzog 1980). Defoliation during pod formation is known to cause a significant reduction in yields (Dungun 1939, Fuellman 1944, Kalton et al. 1945, McAlister and Krober 1958,

Turnipseed 1972). Talekar and Lee (1988), reported that a single defoliation event of 25 or 50% 58

during the R3 to R5 growth stages significantly reduced yields. More recently this threshold level was further refined to 20% defoliation during reproductive growth stages (Owen et al.

2013). Soybeans at R7 (beginning maturity) or later can withstand defoliation since te seeds are mature and are no longer accumulating photosynthate or nutrients (Malone et al. 2002).

Subsequent defoliation events that occur over time through the reproductive growth stages are considered to occur independent of each other (Thrash 2018). Because soybean looper larvae usually begin feeding on foliage about the time that soybeans reach full bloom (R2) in

Mississippi (Fehr et al. 1971), and defoliation at this stage and those that follow it are most critical for yield production, it is important to have updated and accurate thresholds for larval sampling (Carter-Wientjes et al. 2004). At present the threshold for soybean loopers in reproductive stage soybeans is 19 worms 1.27 cm (0.5 in.) or longer per 25 sweeps or four or more worms 1.27 cm (0.5 in.) or longer per row foot (Catchot et al. 2019). Little published data could be found supporting the current larval thresholds and how the current larval thresholds correspond with the defoliation threshold.

Materials and Methods

In 2017 and 2018, maturity group IV soybeans (Asgrow 46X6, Bayer Crop Science, St.

Louis, MO) were planted on 102 cm rows (40” rows) at the Mississippi State University Delta

Research and Extension Center in Stoneville, MS. Plots were arranged in a completely randomized design. In 2017, one trial sampled on 21 Aug and 28 Aug. In 2018, two trials were conducted, both were sampled on 4 Sep and 14 Sep. In each replication one plot was designated as the control plot. Trials were infested with naturally occurring populations of soybean loopers.

Once these populations were observed, the control plots were treated at least weekly until populations declined to prevent establishment and damage from soybean looper larvae. To 59

achieve this, plots were sprayed with either chlorantraniliprole (Prevathon®, FMC, Philadelphia,

PA) or methoxyfenozide + spinetoram (Intrepid Edge®, Corteva, Wilmington, DE). Once feeding occurred, plots were sampled. Each replication was sampled by one individual using 25 sweeps of a 38.1 cm diameter sweep net. Samples were counted and larvae were classified by size.

During 2017, larvae were classified as either less than third instar (less than 1.27 cm (0.5 in.)) or greater than third instar (greater than 1.27 cm (0.5 in.) long). In 2018, larvae were classified as either less than third instar (less than 1.27 cm. (0.5 in.)) or third, fourth, or fifth instar. Once samples were taken, five random plants from each plot were removed and leaf area measure using a Li-Cor 3100C leaf area meter (Li-Cor Biosciences®, Lincoln, NE).

Two sites with naturally occurring soybean looper populations were sampled twice, ten days apart. Larval counts obtained by sweep net sampling were converted to insect feeding equivalents using data from Chapter 3, in an effort to give larger larvae more statistical weight than smaller larvae. Three models were created from this data: conservative, moderate, and aggressive feeding. The insect feeding equivalents from sampling date one and sampling date two were averaged together to account for the cyclical nature of soybean looper populations.

Control plots from each location, across both sampling dates were averaged together to create a more accurate leaf area control to calculate percent defoliation. Defoliation rates were calculated using the equation: % defoliation = ((1 – leaf area of damaged plot / leaf area of control) x 100).

Defoliation rates from sampling date one were subtracted from sampling date two to only consider the defoliation that occurred during the 10-day sampling interval. Although negative defoliation cannot occur naturally, the method used to calculate leaf area, and therefore defoliation, is variable. So as not to skew data, negative defoliation was plotted. The relationship between insect feeding equivalents for all three feeding models from chapter 3 and observed

defoliation rates was determined using regression analysis (PROC GLIMMIX, SAS 9.4, SAS

Institute Inc.) with siteyear and the random term.

Data from the three regression models (conservative, moderate, and aggressive) were then combined with data published by Owen et al. (2013) and Owen (2012) to create prediction charts for various scenarios. In these studies, three economic injury level (percent defoliation) tables were presented based upon the value of the crop ($/ha) and the cost of control ($/ha) of the pest, each of these tables represents the EIL at different growth stages (R3 –Table 4.2, R5 –

Table 4.7 and R6 – Table 4.12). The percent defoliation values were then converted to insect equivalents based on the equations for the regression analysis (Table 4.1). In these equations y represents percent defoliation and x represents insect equivalents. For each EIL table, the percent defoliation was converted into three scenarios of insect feeding (conservative, moderate, and aggressive). Insect equivalents could then be converted to larval counts based on data presented in Tables 3.3 – 3.5. For each model of feeding, for each growth stage, three economic injury levels (larval counts) are proposed based on insect size (third, fourth, and fifth instar larvae).

Results and Discussion

A significant relationship between percent defoliation and the number of soybean looper insect equivalents was observed for the conservative model of feeding (F = 16.19, df = 1,41, P =

0.0002, R2 = 0.3548) (Figure 4.1). A significant relationship between percent defoliation and number of soybean looper insect equivalents was observed for the moderate model of feeding (F

= 15.50, df = 1, 41, P = 0.0003, R2 = 0.3396) (Figure 4.2). A significant relationship between percent defoliation and number of soybean looper insect equivalents was observed for the aggressive model of feeding (F = 14.61, df = 1,41, P = 0.0004, R2 = 0.3298) (Figure 4.3). These regression analyses each represent a different scenario in which defoliation could occur. For 61

example, the aggressive feeding model would represent a situation in which larvae are feeding more aggressively than normal or where plants did not produce an abundant amount of leaf tissue or have significant damage from other defoliators. The moderate model of feeding would represent a situation where soybean looper are feeding normally and plants produce a normal amount of leaf tissue for the variety in question. The conservative feeding model would represent a situation where soybean looper larvae are feeding at a slower or reduced rate or where plants have produced an overabundance of leaf tissue. By using three models of defoliation, three separate economic injury levels can be produced to account for defoliation from other pests at the time of sampling and variability in the leaf tissue growth. Using the equations generated in the regression analysis combined with economic injury level data presented in Owen et al.

(2013) and Owen (2012) a variety of prediction tables could be made based on the growth stage observed and the amount of feeding / leaf area present at the time of sampling. By creating three models of defoliation, three separate economic injury levels can be produced to account for defoliation from other pests at the time of sampling and variability in the leaf tissue growth. For example, if an agricultural professional is sampling a soybean field at R5 growth stage with an average amount of leaf tissue growth, then they would use Table 4.10 to determine the economic injury level of their crop based on the crop value and the cost of control, whereas an agricultural professional sampling a soybean field at the R3 growth stage that has significant defoliation from other pests would use Table 4.6 to determine their economic injury level. After sampling an infested field, the number and growth stage of the larvae should be determined. Since the prediction charts account for larval size, a decision could be made whether to treat or not based on the size and number of larvae present.

In Mississippi in 2018, the average yield was 3,565 kg / ha and the average price was

$0.35 per kilogram making the value of the crop $1,256 / ha and the cost per hectare to treat for soybean looper larvae was $40. Using data from Owen et al. (2013) and Owen (2012) that would make the economic injury level between 18 and 23% defoliation for the R3 growth stage, between 24 and 31% for the R5 growth stage, and between 29 and 35% defoliation for the R6 growth stage. Using Table 4.5, this would indicate that at the R3 growth stage, the economic injury level for larval sampling would be between 22 and 45 third instar larvae, 11 to 21 fourth instar larvae, or 4 to 9 fifth instar larvae. Using Table 4.10 for the R5 growth stage, the economic injury level for larval sampling would be approximately 25 third instar larvae, 12 fourth instar larvae, or 5 fifth instar larvae. Using Table 4.15 for the R6 growth stage, the economic injury level for larval sampling would be approximately 53 third instar larvae, 25 fourth instar larvae, or 10 fifth instar larvae. Taking in consideration that thresholds for larval sampling are set 15 to

25% lower than the economic injury level and the fact that soybean looper infestations are not all going to be at the same growth stage at the same time. The current threshold of 19 larvae 1.27 cm (0.5 in.) or greater per 25 sweeps falls within the acceptable range based on the prediction charts. The constructed models presented in this study do not provide one static number for an economic injury level. When considering an economic injury level, there are numerous factors that must be taken into consideration including: value of the crop, cost of control, overall vigor of the crop, size and density of the infesting population, etc. all of which can impact where an economic threshold is set in relation to an economic injury level. One weakness of these models is that negative or zero (treat at first detection of larvae) economic injury levels were the result of these calculations in some cases, which is unlikely to be biologically or economically feasible.

Therefore, additional research is needed to further refine the models.

Table 4.1 Regression analysis equation for the conservative, moderate, and aggressive feeding models

Feeding Model Regression equation

Conservative Y = 0.0937x + 14.377

Moderate Y = 0.1391x + 14.737

Aggressive Y = 0.161x + 14.178

In above equations, Y = % defoliation and x = number of insect equivalents

Table 4.2 Economic injury levels at R3 and R5 for defoliation based on the yield loss equation for R3 growth stage soybean.

Cost of Control ($/ha)a

Value of Crop $30 $35 $40 $45 $50 $55 ($/ha)

R3 – R5 Economic Injury Level (Percent Defoliation)

2,500 12 13 14 16 18 19

2,000 14 15 16 18 20 22

1,500 16 17 18 20 23 25

1,000 20 21 23 26 29 32

500 28 30 32 36 40 44

Adapted from Owen et al. 2013 a Yields were not significantly different until defoliation exceeded 63% at the R3 and R5 growth stages. All reasonable EIL estimates were below 63%, therefore EILs for the R3 and R5 growth stages were based on the R3 regression equation for defoliation

Table 4.3 Economic injury levels for soybean looper infesting soybean at R3 based on defoliation converted to soybean looper insect equivalents for three feeding models.

Cost of Control ($/ha)

Value of Crop ($/ha) $30 $35 $40 $45 $50 $55

Insect equivalent values from the conservative model

2,500 -25.37a -14.70a -4.02a 17.32 38.67 49.43

2,000 -4.02a 6.65 17.32 38.67 60.01 81.36

1,500 17.32 27.99 38.67 60.01 92.03 113.37

1,000 60.01 70.68 92.03 124.04 156.06 188.08

500 145.39 166.73 188.08 230.77 273.46 316.15

Insect equivalent values from the moderate model

2,500 -13.53a -7.32a -1.11a 11.32 23.74 29.95

2,000 -1.11a 5.11 11.32 23.74 36.16 48.58

1,500 11.32 17.53 23.74 34.16 54.80 67.22

1,000 36.16 42.37 54.80 73.43 92.06 110.70

500 85.85 98.27 110.70 135.54 160.39 185.23

Insect equivalent values from the aggressive model

2,500 -19.68a -12.49a -5.30a 9.08 23.46 30.65

2,000 -5.30a 1.89 9.08 23.46 37.84 52.21

1,500 9.08 16.27 23.46 37.84 59.40 73.78

1,000 37.84 45.03 59.40 80.97 102.54 124.11

500 95.35 109.73 124.11 152.86 181.62 210.37 a Insect equivalents are based on mathematical formulas from regression equations, therefore negative numbers are possible outcomes of the calculations. However, negative insect equivalents are not biologically possible.

Table 4.4 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the conservative feeding model at R3 growth stage

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 2,500 -12.88a -7.46a -2.04a 8.79 19.63 25.04

2,000 -2.04a 3.38 8.79 19.63 30.46 41.30

1,500 8.79 14.21 19.63 30.46 46.71 57.55

1,000 30.46 35.88 46.71 62.97 79.22 95.47

500 73.80 84.64 95.47 117.14 138.81 160.48

Number of fourth instar larvae required to reach EIL

2,500 -2.16a -1.25a -0.34a 1.48 3.30 4.21

2,000 -0.34a 0.57 1.48 3.30 5.12 6.94

1,500 1.48 2.39 3.30 5.12 7.85 9.67

1,000 5.12 6.03 7.85 10.58 13.30 16.03

500 12.39 14.21 16.03 19.67 23.31 26.95

Number of fifth instar larvae required to reach EIL

2,500 -1.16a -0.67a -0.18a 0.79 1.77 2.26

2,000 -0.18 0.30 0.79 1.77 2.75 3.73

1,500 0.79 1.28 1.77 2.75 4.22 5.20

1,000 2.75 3.24 4.22 5.68 7.15 8.62

500 6.66 7.64 8.62 10.58 12.53 14.49 a Insect equivalents are based on mathematical formulas from regression equations, therefore negative numbers are possible outcomes of the calculations. However, negative insect equivalents are not biologically possible.

Table 4.5 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the moderate feeding model at R3 growth stage.

Cost of control ($ / ha) Value of crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 2,500 -5.54a -3.00a -0.45a 4.64 9.73 12.27

2000 -0.45a 2.09 4.64 9.73 14.82 19.91

1,500 4.64 7.18 9.73 14.82 22.46 27.55

1,000 14.82 17.37 22.46 30.09 37.73 45.37

500 35.18 40.28 45.37 55.55 65.73 75.91

Number of fourth instar larvae required to reach EIL

2,500 -2.64a -1.43a -0.22a 2.21 4.64 5.85

2,000 -0.22a 1.00 2.21 4.64 7.06 9.49

1,500 2.21 3.42 4.64 7.06 10.70 13.13

1,000 7.06 8.28 10.70 14.34 17.98 21.62

500 16.77 19.19 21.62 26.47 31.33 36.18

Number of fifth instar larvae required to reach EIL

2,500 -1.06a -0.57a -0.09a 0.88 1.85 2.34

2000 -0.09a 0.40 0.88 1.85 2.82 3.80

1,500 0.88 1.37 1.85 2.82 4.27 5.24

1,000 2.82 3.31 4.27 5.73 7.18 8.63

500 6.70 7.67 8.63 10.57 12.51 14.45 a Insect equivalents are based on mathematical formulas from regression equations, therefore negative numbers are possible outcomes of the calculations. However, negative insect equivalents are not biologically possible.

Table 4.6 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the aggressive feeding model at R3 growth stage.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 2,500 -7.23a -4.59a -1.95a 3.34 8.62 11.27

2,000 -1.95a 0.70 3.34 8.62 13.91 19.20

1,500 3.34 5.98 8.62 13.91 21.84 27.13

1,000 13.91 16.55 21.84 29.77 37.70 45.63

500 35.05 40.34 45.63 56.20 66.77 77.34

Number of fourth instar larvae required to reach EIL

2,500 -3.79a -2.41a -1.02a 1.75 4.52 5.91

2,000 -1.02a 0.36 1.75 4.52 7.29 10.06

1,500 1.75 3.13 4.52 7.29 11.45 14.22

1,000 7.29 8.68 11.45 15.60 19.76 23.91

500 18.37 21.14 23.91 29.45 34.99 40.53

Number of fifth instar larvae required to reach EIL

2,500 -1.31a -0.83a -0.35a 0.60 1.56 2.04

2,000 -0.35a 0.13 0.60 1.56 2.51 3.47

1,500 0.60 1.08 1.56 2.51 3.95 4.90

1,000 2.51 2.99 3.95 5.38 6.81 8.25

500 6.34 7.29 8.25 10.16 12.07 13.98

Table 4.7 Economic injury levels at R3 and R5 for defoliation based on the yield loss equation for R5 growth stage.

Cost of Control ($/ha)a

Value of Crop ($/ha) $30 $35 $40 $45 $50 $55

R3 – R5 Economic Injury Level (Percent Defoliation)

3,000 17 18 18 20 23 25

2,500 18 19 19 21 24 26

1,875 19 20 21 24 26 29

1,250 22 23 24 27 30 33

625 28 30 31 35 39 43

Adapted from Owen 2012 a R3 and R5 yields were not significantly different until defoliation exceeded 63% and all reasonable EIL estimates are below 63%, EILs at R3 and R5 growth stages were based on the R3 regression equation for defoliation.

Table 4.8 Economic injury levels for soybean looper infesting soybean at R5 based on defoliation converted to soybean looper insect equivalents for three feeding models.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Insect equivalents based on the conservative model 3,000 28.00 38.67 38.67 60.01 92.03 113.37

2,500 38.67 49.34 49.34 70.68 102.70 124.04

1,875 49.34 60.01 70.68 102.70 12404 156.06

1,250 81.36 92.03 102.70 134.72 166.73 198.75

625 145.39 166.73 177.41 220.10 262.79 305.47

Insect equivalents based on the moderate model

3,000 16.27 23.46 23.46 37.84 59.40 73.78

2,500 23.46 30.65 30.65 45.03 66.59 80.97

1,875 30.65 37.84 45.03 66.59 80.97 102.54

1,250 52.21 59.40 66.59 88.16 109.73 131.29

625 95.35 109.73 116.92 145.67 174.43 203.18

Insect equivalents based on the aggressive model

3,000 17.53 23.74 23.74 36.16 54.80 67.22

2,500 23.74 29.95 29.95 42.37 61.01 73.43

1,875 29.95 36.16 42.37 61.01 73.43 92.06

1,250 48.58 54.80 61.01 79.64 98.27 116.91

625 85.85 98.27 104.48 129.33 154.17 179.02

Table 4.9 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the conservative feeding model for the R5 growth stage.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 3,000 14.21 19.63 19.63 30.46 46.71 57.55

2,500 19.63 25.04 25.04 35.88 52.13 62.97

1,875 25.04 30.46 35.88 52.13 62.97 79.22

1,250 41.30 46.71 52.13 68.38 84.64 100.89

625 73.80 84.64 90.05 111.72 133.39 155.06

Number of fourth instar larvae required to reach EIL

3,000 2.39 3.30 3.30 5.12 7.85 9.67

2,500 3.30 4.21 4.21 6.03 8.76 10.58

1,875 4.21 5.12 6.03 8.76 10.58 13.30

1,250 6.94 7.85 8.76 11.48 14.21 16.94

625 12.39 14.21 15.12 18.76 22.40 26.04

Number of fifth instar larvae required to reach EIL

3,000 1.28 1.77 1.77 2.75 4.22 5.20

2,500 1.77 2.26 2.26 3.24 4.71 5.68

1,875 2.26 2.75 3.24 4.71 5.68 7.15

1,250 3.73 4.22 4.71 6.17 7.64 9.11

625 6.66 7.64 8.13 10.09 12.04 14.00

Table 4.10 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the moderate feeding model at the R5 growth stage.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 3,000 7.18 9.73 9.73 14.82 22.46 27.55

2,500 9.73 12.27 12.27 17.37 25.00 30.09

1,875 112.27 14.82 17.37 25.00 30.09 37.73

1,250 19.91 22.46 25.00 32.64 40.28 47.91

625 35.18 40.28 42.82 53.00 63.19 73.37

Number of fourth instar larvae required to reach EIL

3,000 3.42 4.64 4.64 7.06 10.70 13.13

2,500 4.64 5.85 5.85 8.28 11.92 14.34

1,875 5.85 7.06 8.28 11.92 14.34 17.98

1,250 9.49 10.70 11.92 15.55 19.19 22.83

625 16.77 19.19 20.41 25.26 30.11 34.96

Number of fifth instar larvae required to reach EIL

3,000 1.37 1.85 1.85 2.82 4.27 5.24

2,500 1.85 2.34 2.34 3.31 4.76 5.73

1,875 2.34 2.82 3.31 4.76 5.73 7.18

1,250 3.79 4.27 4.76 6.21 7.67 9.12

625 6.70 7.67 8.15 10.09 12.03 13.96

Table 4.11 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the aggressive feeding model at the R5 growth stage.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 3,000 5.98 8.62 8.62 13.91 21.84 27.13

2,500 8.62 11.27 11.27 16.55 24.48 29.77

1,875 11.27 13.91 16.55 24.48 29.77 37.70

1,250 19.19 21.84 24.48 32.41 40.34 48.27

625 35.05 40.34 42.98 53.56 64.13 74.70

Number of fourth instar larvae required to reach EIL

3,000 3.13 4.52 4.52 7.29 11.45 14.22

2,500 4.52 5.91 5.91 8.68 12.83 15.60

1,875 5.91 7.29 8.68 12.83 15.60 19.76

1,250 10.06 11.45 12.83 16.99 21.14 25.30

625 18.37 21.14 22.53 28.07 33.61 39.15

Number of fifth instar larvae required to reach EIL

3,000 1.08 1.56 1.56 2.51 3.95 4.90

2,500 1.56 2.04 2.04 2.99 4.42 5.38

1,875 2.04 2.51 2.99 4.42 5.38 6.81

1,250 3.47 3.95 4.42 5.86 7.29 8.72

625 6.34 7.29 7.77 9.68 11.59 13.50

Table 4.12 Economic injury level at R6 for defoliation based on the yield loss equation at R6 growth stage

Cost of Control ($/ha)

Value of Crop ($/ha) $30 $35 $40 $45 $50 $55

R6 economic injury level (percent defoliation)

2,500 19 21 22 25 28 30

2,000 21 23 25 28 31 34

1,500 25 27 29 33 36 40

1,000 30 33 35 39 44 48

500 43 46 50 56 63 69

Adapted from Owen et al. (2013)

Table 4.13 Economic injury levels for soybean looper infesting soybean at R6 based on defoliation converted to soybean looper insect equivalents for three feeding models.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Insect equivalents based on the conservative model 2,500 49.34 70.68 81.36 113.37 145.39 166.73

2,000 70.68 92.03 113.37 145.39 177.41 209.42

1,500 113.37 134.72 156.06 198.75 230.77 273.46

1,000 166.73 198.75 220.10 262.79 316.15 358.84

500 305.47 337.49 380.18 444.22 518.92 582.96

Insect equivalents based on the moderate model

2,500 30.65 45.03 52.21 73.78 95.35 109.73

2,000 45.03 59.40 73.78 95.35 116.92 138.48

1,500 73.78 88.16 102.54 131.29 152.86 181.62

1,000 109.73 131.29 145.67 174.43 210.37 239.13

500 203.18 224.75 253.51 296.64 346.97 390.10

Insect equivalents based on the aggressive model

2,500 29.95 42.37 48.58 67.22 85.85 98.27

2,000 42.37 54.80 67.22 85.85 104.48 123.12

1,500 67.22 79.64 92.06 116.91 135.54 160.39

1,000 98.27 116.91 129.33 154.17 185.23 210.07

500 179.02 197.65 222.50 259.76 303.24 340.51

Table 4.14 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the conservative feeding model at the R6 growth stage.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 2,500 25.04 35.88 41.30 57.55 73.80 84.64

2,000 35.88 46.71 57.55 73.80 90.05 106.30

1,500 57.55 68.38 79.22 100.89 117.14 138.81

1,000 84.64 100.89 111.72 133.39 160.48 182.15

500 155.06 171.32 192.99 225.49 263.41 295.92

Number of fourth instar larvae required to reach EIL

2,500 4.21 6.03 6.94 9.67 12.39 14.21

2,000 6.03 7.85 9.67 12.39 15.12 17.85

1,500 9.67 11.48 13.30 16.94 19.67 23.31

1,000 14.21 16.94 18.76 22.40 26.95 30.59

500 26.04 28.77 32.41 37.87 44.24 49.70

Number of fifth instar larvae required to reach EIL

2,500 2.26 3.24 3.73 5.20 6.66 7.64

2,000 3.24 4.22 5.20 6.66 8.13 9.60

1,500 5.20 6.17 7.15 9.11 10.58 12.53

1,000 7.64 9.11 10.09 12.04 14.49 16.45

500 14.00 15.47 17.42 20.36 23.78 26.72

Table 4.15 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the moderate feeding model at the R6 growth stage.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 2,500 12.27 17.37 19.91 27.55 35.18 40.28

2,000 17.37 22.46 27.55 35.18 42.82 50.46

1,500 27.55 32.64 37.73 47.81 55.55 65.73

1,000 40.28 47.91 53.00 63.19 75.91 86.09

500 73.37 81.00 91.19 106.46 124.28 139.55

Number of fourth instar larvae required to reach EIL

2,500 5.85 8.28 9.49 13.13 16.77 19.19

2,000 8.28 10.70 13.13 16.77 20.41 24.05

1,500 13.13 15.55 17.98 22.83 26.47 31.33

1,000 19.19 22.83 25.26 30.11 36.18 41.03

500 34.96 38.60 43.46 50.74 59.23 66.51

Number of fifth instar larvae required to reach EIL

2,500 2.34 3.31 3.79 5.24 6.70 7.67

2,000 3.31 4.27 5.24 6.70 8.15 9.60

1,500 5.24 6.21 7.18 9.12 10.57 12.51

1,000 7.67 9.12 10.09 12.03 14.45 16.39

500 13.96 15.42 17.36 20.26 23.65 26.56

Table 4.16 Economic injury levels for third, fourth, and fifth instar soybean looper larvae based on insect equivalents for the aggressive feeding model at the R6 growth stage.

Cost of Control ($ / ha)

Value of Crop ($ / ha) $30 $35 $40 $45 $50 $55

Number of third instar larvae required to reach EIL 2,500 11.27 16.55 19.20 27.13 35.05 40.34

2,000 16.55 21.84 27.13 35.05 42.98 50.91

1,500 27.13 32.41 37.70 48.27 56.20 66.77

1,000 40.34 48.27 53.56 64.13 77.34 87.92

500 74.70 82.63 93.20 109.06 127.56 143.42

Number of fourth instar larvae required to reach EIL

2,500 5.91 8.68 10.06 14.22 18.37 21.14

2,000 8.68 11.45 14.22 18.37 22.53 26.68

1,500 14.22 16.99 19.76 25.30 29.45 34.99

1,000 21.14 25.30 28.07 33.61 40.53 46.08

500 39.15 43.30 48.85 57.16 66.85 75.16

Number of fifth instar larvae required to reach EIL

2,500 2.04 2.99 3.47 4.90 6.34 7.29

2,000 2.99 3.95 4.90 6.34 7.77 9.20

1,500 4.90 5.86 6.81 8.72 10.16 12.07

1,000 7.29 8.72 9.68 11.59 13.98 15.89

500 13.50 14.93 16.84 19.71 23.05 25.92

Figure 4.1 Relationship between observed percent defoliation and number of soybean looper insect equivalents calculated based on the conservative model of over a 10-day period.

Figure 4.2 Relationship between observed percent defoliation and number of soybean looper insect equivalents calculated based on the moderate model of feeding over a 10- day period.

Figure 4.3 Relationship between observed percent defoliation and number of soybean looper insect equivalents calculated based on the aggressive model of feeding over a 10- day period.

References Cited

Barrionueva, M. J., M. G. Murua, L. G. R. Meagher, and F. Navarro. 2012. Life table studies of Rachiplusia nu (Guenee) and Chrysodeixis (=Pseudoplusia) includens (Walker) (Lepidoptera: Noctuidae) on artificial diet. Fl. Entomol. 85: 944 – 951.

Carter-Wientjes, C. H., J. S. Russin, D. J. Boethel, J. L. Griffin, and E. C. McGawley. 2004. Feeding and maturation by soybean looper (Lepidoptera: Noctuidae) larvae on soybean affected by weed, fungus, and nematode. J. Econ. Entomol. 97: 14-20. Dungun, G. H. 1939. Recovery of soybeans from hail is measured, pp. 86-87. Ann. Rep. 50. Il Agr. Exp. Stat. Univ. of Illinois. Urbana-Champaign, IL. Fehr, W. R., C. E. Caviness, D. T. Burwood, and J. S. Pennington. 1971. Stages of development descriptions for soybeans, Glycine max (L.) Merrill. Crop Sci. 11: 929-931. Fehr, W. R., Froehlich, D. M., Ertl, D.S. 1985. Iron-deficiency chlorosis of soybean cultivars injured by plant cutoff and defoliation. Crop Sci. 25: 21-23. Fuellman, R.F. 1944. Hail damage to soybeans. Trans. Ill. State Acad. Sci. 37: 25-28. Herbert, Jr., D. A., T. P. Mack, P. A. Backman, and R. Rodrigues-Kabana. 1992. Validation of a model for estimating leaf-feeding by insects in soybeans. Crop Protec. 11: 27-34. Herzog, D. C. 1980. Sampling soybean looper on soybean, pp. 141-168. In M. Kogan and D. C. Herzog (ed.), Sampling methods in Soybean Entomology, 1st ed. Springer-Verlag New York Inc. Kalton, R. R., C. R. Weber, and J. C. Eldridge. 1945. The effect of injury simulation hail damage to soybeans, pp. 733-796. Iowa Agr. Exp. Sta. Iowa State Univ. Ames, IA. Klubertanz, T. H., L. P. Pedigo, and R. E. Carlson. 1996. Reliability of yield models of defoliated soybean based on leaf area index versus leaf area removed. J. Econ. Entomol. 89: 751-756. Malone, S., D. A. Herbert Jr., and D. L. Holshouser. 2002. Relationship between leaf area index and yield in double-crop and full-season soybean systems. J. Econ. Entomol. 95: 945-951. McAlister, D. F. and O. A. Krober. 1958. Response of soybean to leaf and pod removal. Agron. J. 50: 674-677. Musser, F. R., A. L. Catchot, Jr., S. P. Conley, J. A. Davis, C. DiFonzo, J. Greene, G. M. Lorenz, D. Owens, T. Reed, D. D. Reisig, P. Roberts, T. Royer, N. J. Seiter, S. D. Stewart, S. Taylor, K. Tilmon, and M. O. Way. 2018. 2017 Soybean insect losses in the United States. Midsouth Entomol. 11: 1-23. 82

Musser, F. R., A. L. Catchot, Jr., J. A. Davis, G. M. Lorenz, T. Reed, D. D. Reisig, S. D. Stewart, and S. Taylor. 2017. 2016 Soybean insect losses in the Southern US. Midsouth Entomol. 10: 1-13.

Musser, F. R., A. L. Catchot, Jr., J. A. Davis, D. A. Herbert, Jr., G. M. Lorenz, T. Reed, D. D. Reisig, and S. D. Stewart. 2016. 2015 soybean insect losses in the southern US. Midsouth Entomol. 9: 5-17.

Musser, F. R., A. L. Catchot, Jr., J. A. Davis, D. A. Herbert, Jr., G. M. Lorenz, T. Reed, D. D. Reisig, and S. D. Stewart. 2015. 2014 Soybean insect losses in the Southern US. Midsouth Entomol. 8: 35-48.

Musser, F. R., A. L. Catchot, Jr., J. A. Davis, D. A. Herbert, Jr., G. M. Lorenz, T. Reed, D. D. Reisig, and S. D. Stewart. 2014. 2013 Soybean insect losses in the Southern US. Midsouth Entomol. 7: 15-28.

Musser, F. R., A. L. Catchot, Jr., J. A. Davis, D. A. Herbert, Jr., G. M. Lorenz, T. Reed, D. D. Reisig, and S. D. Stewart. 2013. 2012 Soybean insect losses in the Southern US. Midsouth Entomol. 6: 12-24.

Musser, F. R., A. L. Catchot, Jr., J. A. Davis, D. A. Herbert, Jr., B. R. Leonard, G. M. Lorenz, T. Reed, D. D. Reisig, and S. D. Stewart. 2012. 2011 Soybean insect losses in the Southern US. Midsouth Entomol. 5: 11-22.

Musser, F. R., G. M. Lorenz, S. D. Stewart, and A. L. Catchot, Jr. 2011. 2010 Soybean insect losses for Mississippi, Tennessee, and Arkansas. Midsouth Entomol. 4: 22-28.

Musser, F. R., G. M. Lorenz, S. D. Stewart, and A. L. Catchot, Jr. 2010. 2009 Soybean insect losses for Mississippi, Tennessee, and Arkansas. Midsouth Entomol. 3: 48-54.

Musser, F. R., S. D. Stewart, and A. L. Catchot, Jr. 2009. 2008 Soybean insect losses for Mississippi and Tennessee. Midsouth Entomol. 2: 42-46

Musser, F. R., and A. Catchot. 2008. Mississippi soybean insect losses. Midsouth Entomol. 1: 29-36.

Owen, Lucas N. 2012. Effects of defoliation in soybeans and susceptibility of soybean loopers to reduced risk insecticides. Doctor of Philosophy in Life Sciences Dissertation. Mississippi State University. Mississippi State, Mississippi.

Owen, L. N., A. L. Catchot, F. R. Musser, J. Gore, D. Cook, R. Jackson, and C. Allen. 2013. Impact of defoliation on yield on group IV soybean in Mississippi. Crop Prot. 54: 206- 212. Talekar, N. S. and H. R. Lee. 1988. Response of soybean to foliage loss in Taiwan. J. Econ. Entomol. 81: 1363-1368.

Thrash, B. C. 2018. Evaluation of soybean production practices that impact yield losses from simulated insect defoliation. Doctor of Philosophy in Life Sciences, Mississippi State University Mississippi State, Mississippi. Turnipseed, S. G. 1972. Response of soybeans to foliage losses in South Carolina. J. Econ. Entomol. 65: 224-229.

CHAPTER V

SUMMARY OF STUDIES

In Mississippi, soybean looper, Chrysodeixis includens (Walker), usually infests soybean during mid-August through September. During this time, the majority of soybean in Mississippi, and through-out the Mid-southern United States, will be in the reproductive growth stages.

During the reproductive stages, defoliation of 13.3 to 50% can result in yield losses, depending on the vigor and value of the crop (Bidwell 1979, Pickle and Caviness 1984, Browde et al. 1994,

Owen et al. 2013). The magnitude of yield loss from defoliation is greatest during the early to mid – reproductive growth stages and diminishes as plants approach maturity (Owen et al. 2013).

The defoliation threshold of 20% for soybean during the reproductive growth stage has recently been validated and refined for the current production system in Mississippi (Owen et al. 2013).

The recent validation of defoliation thresholds has led to the reevaluation of current larval thresholds (19 larvae greater than or equal to 1.27 cm (0.5 inch) per 25 sweeps; or 20 larvae greater than or equal to 1.27 cm (0.5 inch) or larger per 1.524 row meters (5 row feet)) (Catchot et al. 2019). Studies were conducted to develop a conversion between the drop cloth and sweep net sampling methods. The sweep net method was more effective at capturing larvae both less than third instar and larvae greater than or equal to third instar. Although the majority of agricultural professionals use a sweep net to sample soybean, having these conversions will help refine the drop cloth threshold. Conversions will also be useful in research situations where field cages are used. Often field cages are not large enough for sweep net sampling to occur therefore

data collected using the drop cloth sampling method could be converted to a sweep net threshold for comparison with the sweep net thresholds.

A study was conducted to determine the soybean leaf consumption rate of soybean looper larvae of various developmental stages (instar). Soybean looper larvae less than third instar consumed less soybean leaf area than larvae of other developmental stages (third, fourth, and fifth instar). Third and fourth instar larvae consumed similar amounts of leaf area, while fifth instar larvae consumed significantly more leaf area than larvae of other developmental stages.

Data from the current study were combined with data published in: Reid and Green 1973, Kogan and Cope 1974, Boldt et al. 1975, and Trichillo and Mack 1989. Feeding ranges were then established for each instar, the lowest number from these ranges for each instar was used in the conservative feeding model, the mean number used for the moderate feeding model, and the highest number used for the aggressive feeding model. Each model was then converted to insect equivalents for further use.

Finally, a field study was conducted in which soybean larval density and percent defoliation was determined. Using regression analysis and the three previously mentioned feeding studies, three scenarios were established: one in which larvae fed conservatively or there was an excess of leaf tissue present, one in which larval fed at an average rate and there was an average amount of leaf tissue present, and one in which larvae fed aggressively or there was a limited amount of leaf tissue present. Each of these models was subsequently used in conjunction with economic injury levels based on percent defoliation established in Owen et al. (2013) and

Owen (2012) to create three larval economic injury level tables each for the R3, R5, and R6 growth stage.

The establishment of models that directly relate to the economic injury level charts for the R3, R5 and R6 growth stage (Owen 2012, Owen et al. 2013) provide a way for agricultural professionals to not only determine the defoliation economic injury level for their crop, but also have a related larval density economic injury level that is based on the same value of crop / cost of control equation as the defoliation economic injury level. However, these economic injury levels are derived using mathematical formulas, and in some cases the results were negative or zero (treat at first detection of larvae), which is unlikely to be biologically or economically feasible. Therefore, additional research is needed to further refine the models.

References Cited

Bidwell, R. G. S. 1979. Plant physiology. In The Macmillan biology series. 2nd ed. Macmillan. New York, NY.

Browde, J. A., L. P. Pedigo, M. D. K. Owen, G. L. Tylka, B. C. Levene. 1994. Growth of soybean stressed by nematodes, herbicides, and simulated insect defoliation. Agron. J. 86: 968-974.

Owen, L. N., A. L. Catchot, F. R. Musser, J. Gore, D. Cook, R. Jackson, and C. Allen. 2013. Impact of defoliation on yield group IV soybean in Mississippi. Crop Prot. 54: 206-212.

Pickle, C. S., and C. E. Caviness. 1984. Yield reduction from defoliation and plant cutoff of determinate and semi-determinate soybean. Ag. J. 76: 474-476.

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