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EFFECTS OF INTERCROPPING AND BIOLOGICAL INSECTICIDES ON SUGARCANE APHID (HEMIPTERA: APHIDIDAE) INFESTATIONS ON SORGHUM, AND IDENTIFICATION OF NATURAL ENEMIES AND ALTERNATE HOSTS IN HAITI

WILFRID CALVIN

A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE

UNIVERSITY OF FLORIDA

2019

To Jehovah, Issa, Calissa, Amelise, and Mercilhome

ACKNOWLEDGMENTS

I thank God for always holding my hand through every step in my life. I am also grateful to my family for their unfailing support throughout my life. I would like to thank my lovely wife for her undying assistance and constant encouragement during my study period. Special thanks to my adorable daughter who endured with love such a long period of time away from daddy to make this achievement possible.

I thank Dr. Julien Beuzelin, my committee chair, for all his guidance and support during my master’s study. My committee members, Drs. Oscar Liburd and Marc

Branham, have also provided useful advice and support for which I am so thankful. I am also thankful to Mr. Ludger Jean Simon for his support toward the success of the experiments conducted in Haiti. I would like to thank Dr. Elijah Talamas for his help identifying insect samples from Haiti.

I thank Donna Larsen for providing technical assistance in all experiments conducted at the UF/IFAS Everglades Research and Education Center (EREC) and for all the help to make my stay in Belle Glade successful. I am also thankful to Erik Roldán for all his help during my master’s program. Blanc Sylvain, Ludger Roland, Joab Calvin,

Jean Benito Chery, Anne Metushelah Eliscar, Rochelin Zakary, Wikenson Dérival, and

Amir Avila provided technical assistance for which I am so grateful.

I thank USAID project Feed the Future Haiti / Appui à la Recherche et au

Développement Agricole (AREA) for granting me the opportunity to pursue this Master of Science degree at the University of Florida. I am also thankful to all the EREC staff, faculty, and students for all their supports without which my time in Belle glade would have been unbearable. My Haitian friends at the University of Florida have helped me to make this journey possible and I will always be grateful.

TABLE OF CONTENTS

page

ACKNOWLEDGMENTS ...... 4

LIST OF TABLES ...... 7

LIST OF FIGURES ...... 8

ABSTRACT ...... 9

CHAPTER

1 INTRODUCTION ...... 11

Importance of Sorghum in Haiti ...... 11 Pest Status of the Sugarcane Aphid on Sorghum ...... 11 Sugarcane Aphid Biology ...... 12 Sugarcane Aphid Predators and Parasitoids ...... 13 Entomopathogens and Potential Role in Sugarcane Aphid Management...... 14 Conventional Insecticides for Sugarcane Aphid Management ...... 17 Botanical Insecticides ...... 17 Host Plant Resistance and Cultural Practices ...... 19 Research Objectives ...... 21

2 EFFECTS OF SORGHUM INTERCROPPING WITH MAIZE AND PIGEON PEA ON SUGARCANE APHID (HEMIPTERA: APHIDIDAE) INFESTATIONS ON SORGHUM AND IDENTIFICATION OF SUGARCANE APHID NATURAL ENEMIES AND ALTERNATE HOSTS IN HAITI ...... 22

Materials and Methods...... 25 Experimental Field Plots ...... 25 Sugarcane Aphid Sampling on Sorghum ...... 26 Sorghum Growth and Yield Determination ...... 26 Sugarcane Aphid Natural Enemy Sampling ...... 27 Sugarcane Aphid Alternate Host Sampling ...... 28 Statistical Analysis ...... 29 Results ...... 30 Sugarcane Aphid Infestations on Sorghum ...... 30 Sorghum Growth and Yield ...... 31 Sugarcane Aphid Natural Enemies ...... 32 Sugarcane Aphid Alternate Hosts ...... 32 Discussion ...... 33

3 EFFECTS OF BIOLOGICAL INSECTICIDES ON SUGARCANE APHID (HEMIPTERA: APHIDIDAE) INFESTATIONS ON SORGHUM ...... 42

Materials and Methods...... 45 Experimental Treatments ...... 45 Aphid Colony ...... 46 Laboratory Experiment ...... 47 Greenhouse Experiments ...... 48 Field Experiments ...... 49 Statistical Analysis ...... 50 Results ...... 51 Laboratory Experiment ...... 51 Greenhouse Experiments ...... 52 Field Experiments ...... 52 Discussion ...... 53

4 SUMMARY ...... 63

LIST OF REFERENCES ...... 66

BIOGRAPHICAL SKETCH ...... 77

LIST OF TABLES

Table page

2-1 Cumulative aphid days (CAD) for sugarcane aphids infesting sorghum in sorghum, sorghum-maize, and sorghum-pigeon pea plots, Jonc-Labeille, Haiti, October-December 2018...... 37

2-2 Cumulative aphid days (CAD) for sugarcane aphids infesting sorghum in sorghum, sorghum-maize, and sorghum-pigeon pea plots, Jonc-Labeille, Haiti, March-May 2019...... 38

2-3 Sorghum plant height and stem diameter comparisons in intercropping experiments, Jonc-Labeille, Haiti, 2018 and 2019...... 39

2-4 Sorghum yield comparisons in intercropping experiments, Jonc-Labeille, Haiti, 2018 and 2019...... 40

2-5 Mean number of insects collected with a sweep net in intercropping experiments, Jonc-Labeille, Haiti, 2018 and 2019...... 40

2-6 Number of sugarcane aphids found on itchgrass, Jonc-Labeille, Haiti, 2018 and 2019...... 41

3-1 Sugarcane aphid infestation levels on sorghum plants as affected by the application of biological insecticides and vetiver oil in the greenhouse, Belle Glade, FL, summer 2018...... 59

3-2 Sugarcane aphid infestation levels as affected by insecticide treatment and post-treatment observation date in three field experiments, Belle Glade, FL, 2018 and 2019...... 61

LIST OF FIGURES

Figure page

2-1 Sorghum plant height and stem diameter (LS means) as affected by intercropping treatment, Jonc-Labeille, Haiti, 2018...... 37

2-2 Sorghum plant height and stem diameter (LS means) as affected by intercropping treatment, Jonc-Labeille, Haiti, 2019...... 38

3-1 Sugarcane aphid mortality over time as affected by the application of seven biological insecticides, vetiver oil, and a conventional insecticide under laboratory conditions, summer 2018, Belle Glade, FL, summer 2018...... 59

3-2 Sugarcane aphid infestation levels in a sorghum field experiment evaluating seven biological insecticides at three post-treatment observation dates, spring 2019, Belle Glade, FL, summer 2019...... 62

Abstract of Thesis Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Master of Science

EFFECTS OF INTERCROPPING AND BIOLOGICAL INSECTICIDES ON SUGARCANE APHID (HEMIPTERA: APHIDIDAE) INFESTATIONS ON SORGHUM, AND IDENTIFICATION OF NATURAL ENEMIES AND ALTERNATE HOSTS IN HAITI

Wilfrid Calvin

August 2019

Chair: Julien Beuzelin Major: Entomology and Nematology

The sugarcane aphid (Melanaphis sacchari) emerged as a sorghum (Sorghum bicolor) pest in North America in 2013 and became a concern in sorghum production in

Haiti in 2015. Field experiments were conducted in Jonc-Labeille, Haiti in 2018 and

2019 to determine the effects of intercropping sorghum with maize (Zea mays) or pigeon pea (Cajanus cajan) compared to a sorghum monoculture on sugarcane aphid infestations. In both years, infestation levels were substantially lower in sorghum-maize than in sorghum-pigeon pea or sorghum alone. Relatively small sorghum plant size due to competition with maize plants likely contributed to the decrease in sugarcane aphid infestations in sorghum-maize. The experimental plots were sampled for potential natural enemies. Coccinellids, chrysopids, syrphids, and braconids were observed. All grass species present in the experimental plots and their direct vicinity were inspected to identify potential alternate hosts of the aphid. Itchgrass (Rottboellia cochinchinensis) was observed infested with one to four aphids per plant when infested. Laboratory, greenhouse, and field experiments were conducted in Belle Glade, FL to determine the effects of biological insecticides on sugarcane aphid infestations. Azadirachtin,

pyrethrins, Beauveria bassiana strain GHA, Isaria fumosorosea Apopka strain 97,

Chromobacterium subtsugae strain PRAA4-1T, Burkholderia spp. strain A396, and vetiver oil were compared to a conventional insecticide, flupyradifurone.

Flupyradifurone, azadirachtin, pyrethrins, B. bassiana, C. subtsugae and vetiver oil negatively affected the insect in the laboratory and greenhouse. However, decreases in infestations relative to the non-treated control were not observed in the field although flupyradifurone was consistently associated with the lowest infestations.

CHAPTER 1 INTRODUCTION

Importance of Sorghum in Haiti

Sorghum [Sorghum bicolor (L.) Moench] is the third most important cereal crop in terms of production in Haiti, with 75,000 metric tons in 2016 compared with 240,000 metric tons for maize (Zea mays L.) and 125,000 metric tons for rice (Oryza sativa L.)

(FAO 2017). Sorghum in Haiti is primarily used for human consumption. It is also used as animal feed (Eckert et al. 2017) and as raw product in the production of alcoholic and malted beverages (Leclerc et al. 2013). On land where other crops cannot be grown, farmers rely on sorghum production because the crop is drought tolerant (Kumar 2016) and can be produced under low to no input conditions (Muleta et al. 2019).

Approximately 200,000 Haitian farmers rely on sorghum for income and family consumption (K-State 2016).

Pest Status of the Sugarcane Aphid on Sorghum

The sugarcane aphid [Melanaphis sacchari (Zehntner)] was reported on sugarcane (Saccharum spp. hybrids) in Hawaii in 1895, in Florida in 1977, and in

Louisiana in 1999 (Hall 1987, White et al. 2001, Villanueva et al. 2014). In 2013, this aphid became a major pest of sorghum in North America. By the end of 2015, the sugarcane aphid had caused economically damaging infestations in sorghum fields in

17 states in the United States (Brewer et al. 2016). The same year, this insect became a major pest on sorghum in Haiti (ayiboPost 2017). This sugarcane aphid outbreak caused losses close to 70% in Haiti, which represents more than 60,000 tons of sorghum grain (Gabriel 2016, ayiboPost 2017).

In a study conducted before the outbreaks on sorghum in North America and

Haiti, sugarcane aphids collected on sugarcane, sorghum, and two additional host plants in 14 countries including China, Australia, Mauritius, Benin, Kenya, Guadeloupe,

Martinique, Brazil, and the United States (Hawaii and Louisiana) exhibited low genetic variability (Nibouche et al. 2014). However, some sugarcane aphid lineages have evolved to preferentially feed on plants in the sorghum genus despite the existence of low genetic variability (Nibouche et al. 2015). The sugarcane aphid populations infesting sorghum in North America since 2013 are genetically distinct from populations previously studied in the region, and it has been hypothesized that the outbreaks in sorghum are the result of the introduction of a sugarcane aphid genotype likely from

Asia (Nibouche et al. 2018).

Sugarcane Aphid Biology

Sugarcane aphid individuals can be alate or apterous. They can be light yellow, yellow brown, purple, or pink (Blackman and Eastop 1984). They can also be gray under cool temperatures. Sugarcane aphids have short and dark cornicles, dark tarsi, and thread-like antennae that are darker near the tip (Villanueva et al. 2014, Brewer et al. 2016). Alate aphids have dorsal markings that are dark and sclerotized. Alate and apterous aphids do not differ in size, ranging from 1.1 to 2.0 mm (Blackman and Eastop

1984). Sugarcane aphid nymphs attain adulthood in about 4 days with a life span of up to 4 weeks. Populations of the sugarcane aphid can increase ten-fold in approximately

15 days (Knutson et al. 2015). Aphids are born pregnant and one female is capable of giving birth to approximately 30-60 nymphs during its lifetime (Brown et al. 2015).

Variation in temperature and the amount of rainfall can impact sugarcane aphid populations (Singh et al. 2004). It is likely that all sugarcane aphids are anholocyclic

(i.e., they reproduce parthenogenetically) (Blackman and Eastop 1984) although rare examples of sexual reproduction have been reported (Singh et al. 2004).

The sugarcane aphid host range is limited to plants in the Poaceae family

(Blackman and Eastop 1984). In addition to sorghum, sugarcane aphid crop hosts include rice, maize, and sugarcane. The sugarcane aphid can also be found on weedy grasses including bermudagrass [Cynodon dactylon (L.) Pers.], chinese silvergrass

(Miscanthus sinensis Andersson), jungle rice [Echinochloa colona (L.) Link], guineagrass (Panicum maximum Jacq.), hairy crabgrass [Digitaria sanguinalis (L.)

Scop.], elephant grass (Pennisetum sp.), foxtail millet [Setaria italic (L.) Beauv], johnsongrass [Sorghum halepense (L.) Pers.], and wild sudangrass [Sorghum verticiliflorum (Steud.) Stapf.] (Singh et al. 2004). However, the sugarcane aphid exhibits preference for plants in the genus Sorghum and Saccharum (Blackman and

Eastop 1984, Singh et al. 2004, Nibouche et al. 2015).

Sugarcane Aphid Predators and Parasitoids

Arthropod natural enemies provide significant ecosystem services that favor suppression of agricultural insect pest populations (Safarzoda et al. 2014). The sugarcane aphid alone has more than 47 species of natural enemies worldwide (Singh et al 2004). These natural enemies include Orius insidiosus Say (Hemiptera:

Anthocoridae), Brachiacantha decora Casey (Coleoptera: Coccinellidae), Coccinella septempunctata (L.) (Coleoptera: Coccinelidae), Coleomegilla maculata DeGeer

(Coleoptera: Coccinellidae), Collops vittatus (say) (Coleoptera: Melyridae), Cycloneda sanguinea (L.) (Coleoptera: Coccinellidae), Diomus roseicollis (Mulsant) (Coleoptera:

Coccinellidae), Diomus terminatus (Say) (Coleoptera: Coccinellidae), Harmonia axyridis

Pallas (Coleoptera: Coccinellidae), Hippodamia convergens Guerin-Meneville

(Coleoptera: Coccinellidae), Hyperaspis wickhami Casey (Coleoptera: Coccinellidae),

Olla v-nigrum (Mulsant) (Coleoptera: Coccinellidae), Scymnus (Pullus) loewii Mulsant

(Coleoptera: Coccinellidae), Ceraeochrysa valida (Banks) (Neuroptera: Chrysopidae),

Chrysoperla carnea Stephens (Neuroptera: Chrysopidae), Aphelinus spp. Dalman

(Hymenoptera: Aphelinidae), Lysiphlebus testaceipes (Cresson) (Hymenoptera:

Braconidae), Allograpta obliqua Say (Diptera: Syrphidae), and Trombidium holosericeum (Trombidiformes: Trombidiidae) (Colares et al. 2015a, 2015b, Rodriguez-

Velez et al. 2016, Rodriguez-del-Bosque et al. 2018).

On sugarcane in Florida, a reduction of 5 to 48% of sugarcane aphid populations by natural enemies from early to late summer was observed (Hall 1987). Bowling et al.

(2016) stated that natural enemies alone are not capable of maintaining sugarcane aphid populations under economically damaging levels. However, in an experiment conducted on sorghum infested with sugarcane aphids, Colares et al. (2015a) found that natural enemies suppressed the aphids before they could damage the plants. In

Mexico, a significant decrease in sugarcane aphid abundance and sorghum damage was observed 4 years after the first outbreaks due to natural enemies. The braconid L. testaceipes plays an important role by emerging at the beginning of sugarcane aphid infestations (Rodriguez-del-Bosque et al. 2018).

Entomopathogens and Potential Role in Sugarcane Aphid Management

Entomopathogens such as fungi, bacteria, viruses, and protozoans have pathogenic effects on arthropods and numerous species are suitable for insect pest management (Ramanujam et al. 2014). More than 750 fungi species are insect pathogens (Sheepmaker and Butt 2010). Entomopathogenic fungi attack 20 out of 31

insect orders and can have effects on all developmental stages, from eggs to adults

(Araujo and Hughes 2016). Fungi from the hypomycetes group cause insect death by causing nutritional deficiency, as well as destruction of tissues and release of toxins

(Ramanujam et al. 2014). Fungal spores adhere to the insect host cuticle, then germinate and enter the cuticle, which causes subsequent infection (Roberts 1989).

Roberts (1989) stated that fungi have a particular importance when managing pests that suck plant sap because these insects have no means of ingesting pathogens.

Beauveria bassiana (Bals. -Criv.) Vuill., Beauveria brongniartii (Sacc.) Petch,

Metarhizium anisopliae (Metchikoff) Sorokin, Lecanicillium spp., Hirsutella thompsonii

Fisher, Nomuraea rileyi (Farlow) Samson, and Isaria fumosorosea (Wize) Brown &

Smith are entomopathogenic fungi that have been extensively investigated (Ramanujam et al. 2014).

In a laboratory experiment conducted by Maketon et al. (2013), B. bassiana

CKB-048 sprayed on sugarcane aphid in sorghum was capable of killing up to 90% of sugarcane aphid nymphs. Mixing B. bassiana with other insecticides is a suitable practice that increases its efficacy (Reddy and Antwi 2016). The effects of this fungus have also been studied on numerous other insect pests. Laboratory bioassay studies showed that the application of a high concentration (1x108 spores/ml) of B. bassiana caused >96% mortality in the cowpea aphid [Aphis craccivora (Koch)] (Saranya et al.

2010).

To the best of our knowledge, the effect of Isaria fumosorosea (Wize) Brown &

Smith against the sugarcane aphid has not been studied. Nonetheless, the effect of this fungus has been investigated on other insect pests such as the red palm weevil

(Rhynchophorus ferrugineus Olivier), beet armyworm [Spodoptera exigua (Hübner)], fall armyworm [Spodoptera frugiperda (J.E. Smith)], and sweet potato whitefly [Bemisia tabaci (Gennadius)] (Sabbour and Abdel-Raheem 2014). Isaria fumosorosea can be used to manage red palm weevils because it can cause up to 100% egg and larval mortality (Sabbour and Abdel-Raheem 2014). Gandarilla-Pacheco et al. (2015) reported that immersing neonate beet armyworms and corn earworms [Helicoverpa zea

(Boddie)] in a solution of I. fumosorosea isolates at a concentration of 1x108 conidia/ml killed 40 and 57% of neonates, respectively. Spraying using microdroplet applicators

(droplets 1-30 and 30-90 µm) caused 70 to 90 % larval mortality (Gandarilla-Pacheco et al. 2015).

Chromobacterium subtsugae strain PRAA4-1T registered as Grandevo (Marrone

Bio Innovations, Davis, CA) in the United States has been tested on the sugarcane aphid. One application of Grandevo did not cause significant effects on infestations on sorghum in one field study (Studebaker and Jackson 2017). In addition, Natwick and

Lopez (2015) did not observe measurable effects against the pea aphid (Acyrthosiphon pisum Harris) in alfalfa (Medicago sativa L.). However, in an evaluation conducted on hairy chinch bugs (Blissus leucopterus hirtus Montandon) in bluegrass (Poa spp.), different control levels were observed using Grandevo depending on the number of days after treatment and product rate. A rate of 12.2 kg/ha provided 81.8, 72.6 and

70.8% control at 7, 14, and 29 days after treatment, respectively, whereas a rate of 2.2 kg/ha provided 39.8, 82.1, and 36.7% control 7, 14, and 29 days after treatment, respectively (Andon and Shetlar 2015). Grandevo can also decrease melon aphid

(Aphis gossypii Glover) infestations (Kuhar and Doughty 2016). In addition, C.

subtsugae has been reported to cause black pecan aphid [Melanocallis caryaefoliae

(Davis)] mortality up to 96%, providing 55% damage reduction within 3 days (Shapiro-

Ilan et al. 2013).

Burkholderia spp. strain A396 registered as Venerate (Marrone Bio Innovations,

Davis, CA) in the United States has not controlled the sugarcane aphid infesting sorghum in one field evaluation (Studebaker and Jackson 2017). Nevertheless,

Venerate provided significant control for the cranberry fruitworm [Acrobasis vaccinia

(Riley)] in blueberry (Vaccinium spp.) (Wise et al. 2015).

Conventional Insecticides for Sugarcane Aphid Management

Foliar applications of sulfoxaflor (Transform 50 WG, Dow Agrosciences,

Indianapolis, IN) and flupyradifurone (Sivanto, Bayer Crop Science, Research Triangle

Park, NC) can be used to effectively treat sorghum fields infested by the sugarcane aphid with limited negative impacts on beneficial insects (Bowling et al. 2016, Brown et al. 2015). Chlorpyrifos (e.g., Lorsban, Dow Agrosciences) can also be used despite relatively lower efficacy and residual activity. Additionally, neonicotinoid insecticide seed treatments (imidacloprid, thiamethoxam, clothianidin) protect seedlings from infestations for as long as 40 days after planting (Brown et al. 2015). Sulfoxaflor, flupyradifurone, and neonicotinoids all target the nicotinic acetylcholine receptor (IRAC sub-group 4C,

4D, and 4A, respectively) whereas chlorpyrifos inhibits acetylcholine esterase (IRAC sub-group 1B) (IRAC 2019). Thus, the number of effective insecticidal modes of action against the sugarcane aphid is limited.

Botanical Insecticides

Products extracted from seeds of the neem tree (Azadirachta indica A. Juss)

have been used as botanical insecticides for more than 30 years, including in Haiti

(Timyan 1996, Kraiss and Cullen 2008). In neem seeds, nearly 99 biologically active compounds are found with azadirachtin, nimbin, nimbidin, and nimbolides as key molecules (Dua et al. 2009). A commercial neem-based insecticide, Margosan-O, is lethal to the pea aphid by affecting its molting process, reducing its life span and fecundity (Stark and Walter 1995). Eggs from adult aphids treated with azadiractin may have chorion imperfections resulting in reduced viability and susceptibility to infections by fungi. Viviparous aphid progenies are also impacted by azadiractin (Lowery and

Isman 1994).

Pyrethrins, which are compounds naturally occurring in Chrysanthemum plants, have also long been used to control numerous insect pests of crops and livestock

(Casida 1980). There are six insecticidal esters that are referred to as pyrethrins in the

Chrysanthemum plant extract known as pyrethrum. They contain acid and alcohol groups that are substitutes of cyclopropanecarboxylic acid and cyclopentenolone, respectively (Casida 1980). Pyrethroids, which are synthetic pyrethrins, impair voltage- gated sodium channels in nerve cell membranes and disrupt nerve impulse transmission. This condition subsequently leads to paralysis and death (Davies et al.

2007).

Products extracted from vetiver grass (Chrysopogon zizanioides (L.) Roberty) roots have been used as bioinsecticides and insect repellents (Zhu et al. 2003). Vetiver oil is composed of numerous components, including -vetivones, -vetivones, khusinol, khusilal, diethyl phthalate, vetiselinol, khusimol, isovelencenol, and vetivenic acid. Oil composition can vary with vetiver origins. This oil is an effective repellent of Formosan

subterranean termites (Coptotermes formosanus Shiraki.) Most Haitian vetiver oils tested on Formosan termites have shown high repellency capacity plausibly due to the presence of - vetivones, -vetivones, and khusimol (Zhu et al. 2003). Vetiver oil also repels cockroaches and flies (Jain et al. 1982). Whole vetiver grass plants can also negatively impact insects. The spotted stem borer [Chilo partellus (Swinhoe)], which is a pest of grass crops, prefers to lay eggs on vetiver grass, a host that decreases survival of borer offspring (Van den Berg et al. 2003).

Host Plant Resistance and Cultural Practices

Brown et al. (2015) indicated that the use of resistant varieties is essential to manage the sugarcane aphid in sorghum. Sorghum varieties resistant to the insect in the United States include SC110, SC170, Tx2783, Tx3408, Tx3409, B11070, AB11055-

WF1-CS1/RTx436 and AB11055-WF1-CS1-/RTx437 (Bowling et al. 2016). Sorghum can exhibit antibiosis, antixenosis, and tolerance to the sugarcane aphid (Sharma et al.

2013, 2014, Armstrong et al. 2015). In a study including 31 sorghum genotypes,

Sharma et al. (2013) found 21 genotypes that were considerably less affected by the sugarcane aphid, including ICSV 12001, ICSV 12005, IS 21808, and ICSV 745.

Although infestation levels were high, these genotypes exhibited low damage, indicating that they had some level of tolerance to the sugarcane aphid. The genotypes RSV

1211, RS 29, RSV 1338, EC 8-2, PU 10-1, IS 40617, and ICSB 695 are also tolerant to the sugarcane aphid (Sharma et al. 2014). The ability of sorghum to grow under high aphid infestations is also an indicator of tolerance. For instance, RTx2783, a sorghum line highly tolerant to the sugarcane aphid could produce up to 7 leaves per plant under aphid infestations (Armstrong et al. 2015). The genotypes ICSB 323, ICSB 215, ICSB

12004, ICSR 165, IS 40615, ICSV 12001, ICSB 321, and ICSB 724 express antibiosis, limiting the rate of increase of sugarcane aphids (Sharma et al. (2013). The genotypes

B11055, R13219, and RTx2783 express antixenosis, allowing very few nymphs to become established on plants (Armstrong et al. 2017).

Planting sorghum early can help avoid sugarcane aphid infestations in the southern United States (Brown et al. 2015). In addition, high plant density reduces sorghum vigor, which reduces sugarcane aphid numbers and may be beneficial (Singh et al. 2004). Destroying sorghum crop residues and eliminating alternate hosts are recommended cultural methods that reduce pest abundance (Singh et al. 2004). An appropriate crop fertilization should also be maintained because excess in nitrogen can correlate with an aphid population increase (Lama et al. 2019).

Intercropping is a suitable agricultural practice for insect pest management (Khan

2000). Landscape diversity plays an important role in biodiversity conservation and sustainable pest management (Altieri 1999, Bianchi et al. 2006, Rundlof and Smith

2006). Crops grown in intercropping systems are more likely to be less injured than those grown in monoculture. One reason is that the intercrops promote an environment that attracts natural enemies of the pests (Trenbath 1993). Non-host plants can have repellent or deterrent properties that act against the target insects. They can also reduce insect pest ability to locate hosts (Cook et al. 2007) and increase natural enemy abundance (Abate et al. 2000, Cook et al. 2007). For instance, experiments comparing sorghum intercropped with greenleaf desmodium [Desmodium intortum (Mill.) Urb.] and sorghum in monoculture showed that intercropped sorghum was less injured by the stem borer C. partellus and yielded more than sorghum in monoculture (Khan 2006). In

addition, maize intercropped with legumes or with cassava (Manihot esculenta Crantz) reduced the number of maize ear borer (Mussidia nigrivenella Ragonot) eggs by more than 25% and larvae by up to 53%. Maize yield losses were also reduced by 47 to 84%

(Agboka et al. 2006).

Research Objectives

Sugarcane aphid management in Haiti has focused on the development and propagation of sugarcane aphid resistant sorghum varieties (ayiboPost 2017, Flecher

2017). Nonetheless, a combination of effective sugarcane aphid management tactics in

Haiti has yet to be identified. Thus, there is a need to develop an integrated pest management (IPM) strategy that reduces the need for insecticides and is sustainable in the long-term. Reducing the need for insecticides in sugarcane aphid management is especially important because the majority of Haitian famers are smallholders cultivating very small plots generally less than one hectare. Fields are often located near homes and water sources. In addition, the farmers are not well equipped and trained to use pesticides (USAID 2010). The objectives of this research are to:

1. Evaluate the effects of maize and pigeon pea intercropping with sorghum on

sugarcane aphid infestations on sorghum

2. Identify potential natural enemies and alternate host plants of the sugarcane

aphid in Haiti

3. Evaluate the efficacy of seven biological insecticides registered in the United

States, as well as vetiver oil, for control of the sugarcane aphid in the laboratory,

greenhouse, and field

CHAPTER 2 EFFECTS OF SORGHUM INTERCROPPING WITH MAIZE AND PIGEON PEA ON SUGARCANE APHID (HEMIPTERA: APHIDIDAE) INFESTATIONS ON SORGHUM AND IDENTIFICATION OF SUGARCANE APHID NATURAL ENEMIES AND ALTERNATE HOSTS IN HAITI

The sugarcane aphid [Melanaphis sacchari (Zehntner)] emerged as a sorghum

[Sorghum bicolor (L.) Moench] pest in North America in 2013 (Villanueva et al. 2014) and became a concern in sorghum production in Haiti in 2015. The 2015 aphid outbreak in Haiti caused losses approaching 70%, which represents more than 60,000 tons of sorghum grain (Gabriel 2016, ayiboPost 2017). Sorghum is the third most valuable cereal crop in Haiti, with approximately 200,000 Haitian farmers relying on this crop for income and food (K-state 2016, FAO 2017). In addition to its use as food, sorghum grain is used as animal feed and raw product in the alcoholic and malted beverage industries (Leclerc et al. 2013, Eckert et al. 2017). As a drought tolerant and low input crop, sorghum is well adapted to Haitian farming (Kumar 2016, Muleta et al. 2019).

Thus, the sugarcane aphid is a severe threat to a major agricultural commodity in Haiti.

The development of resistant sorghum varieties has been the main approach for sugarcane aphid management in Haiti (ayiboPost 2017, Flecher 2017). Insecticides have also been used by a limited number of sorghum farmers. However, reducing the need for insecticides in sugarcane aphid management is important because the majority of Haitian famers are smallholders cultivating plots generally less than one hectare.

Agricultural fields are often located near homes and water sources. In addition, farmers are not well equipped and trained to use pesticides (USAID 2010). Thus, the adoption of appropriate cultural practices and conservation of natural enemies complementing other sugarcane aphid management tactics might assist in developing a sustainable sugarcane aphid management strategy in Haiti.

More than 75% of sorghum fields in Haiti are managed under an intercropping system (Leclerc et al. 2013, Eckert et al. 2017). Intercropping sorghum with maize (Zea mays L.) or pigeon pea [aka Congo beans, Cajanus cajan (L.) Millsp.] is common (Wall and Meckenstock 1992, Leclerc et al. 2013). In these intercropping systems, sorghum is generally planted 4 weeks after maize or pigeon pea (Levesque 2014), which might affect sugarcane aphid infestations. Intercropping has been successful against numerous insect pests, including the spotted stem borer [Chilo partellus (Swinhoe)] in sorghum (Khan 2006) and maize ear borer (Mussidia nigrivenella Ragonot) in maize

(Agboka et al. 2006). The companion crop can serve as a repellent, decrease visual stimuli, or trap the pest, and can also recruit pest natural enemies by providing more shelter and food sources for predators and parasitoids (Smith and McSorley 2000,

VanTine and Verlinden 2003, Jones and Gillett 2005, Rodriguez-Saona 2012).

Beneficial arthropods provide ecosystem services that can favor suitable control of insect pests (Sarfarzoda et al. 2014). Thirteen lady beetle species (Coleoptera:

Coccinellidae), three hoverfly species (Diptera: Syrphidae), two lacewing species

(Neuroptera: Chrysopidae), one minute pirate bug species (Hemiptera: Anthocoridae), one mite species (Trombidiformes: Trombidiidae), eight braconid species

(Hymenoptera: Braconidae), and one aphelinid species (Hymenoptera: Aphelinidae) have been described as sugarcane aphid natural enemies in Mexico and the United

States (Colares et al. 2015a, 2015b, Bowling et al. 2016, Salas-Araiza et al. 2017,

Rodriguez-del-Bosque et al. 2018, Rodriguez-Velez et al. 2016, 2019). In Haiti, one species of parasitoid (Hymenoptera: Braconidae), two species of lady beetle

(Coleoptera: Coccinellidae), and one species of hoverfly (Diptera: Syrphidae) were also

reported attacking cotton aphid (Aphis gossypii Glover) infestations on cotton

(Gossypium hirsutum L.) and palay rubbervine [Cryptostegia grandiflora (Roxb. Ex R.

Br.) R. Br.] (Knight 1944).

Lysiphlebus testaceipes Cresson (Hymenoptera: Braconidae) can potentially control the sugarcane aphid when occurring at the beginning of an infestation

(Rodriguez-del-Bosque et al. 2018). Predators and parasitoids including Allograta obliqua (Say) (Diptera: Syrphidae), Hippodamia convergens (Guerin-Meneville)

(Coleoptera: Coccinellidae), Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae), and Aphelinus sp. Dalman (Hymenoptera: Aphelinidae) can completely control sugarcane aphid infestations on potted sorghum (Colares et al. 2015a).

The sugarcane aphid has been reported to utilize several weedy grasses including johnsongrass [Sorghum halepense (L.) Pers.], bermudagrass [Cynodon dactylon (L.) Pers.], jungle rice [Echinochloa colona (L.) Link], and guineagrass

(Panicum maximum Jacq.) as an alternate host to sorghum. These alternate hosts, if present, can shelter sugarcane aphid and subsequently become sugarcane aphid reservoirs for future sorghum infestations (Vedrine 2003, Singh et al. 2004, Candy

2008, M. D. Dorval, personal communication, W. Calvin, personal observation).

In this study the effects of maize and pigeon pea intercropping with sorghum on sugarcane aphid infestations on sorghum were evaluated. In addition, potential natural enemies of sugarcane aphid in sorghum in Haiti were identified. Potential sugarcane aphid alternate hosts were also sampled.

Materials and Methods

Experimental Field Plots

Two field experiments were conducted in 2018 and 2019 at Jonc-Labeille

(18.218973° N, 73.884286° W), a locality in southern Haiti. In each experiment, there were two intercropped treatments, sorghum-maize and sorghum-pigeon pea, and a non-intercropped treatment with sorghum only. The local sorghum variety “Bout gode” was used. “Bout gode” is grown as a grain crop in Haiti and is highly susceptible to the sugarcane aphid. The maize variety “Chicken corn” and pigeon pea variety “Pwa kongo local”, which are traditionally grown in Haiti, were used.

Treatments were assigned to plots 6 m long and 8 rows wide (70-cm row spacing) following a randomized complete block design with six blocks (1 replication per block). Maize and pigeon pea were planted on August 22 in 2018 and on January 19 in

2019. Sorghum was planted 4 weeks after maize and pigeon pea on September 19 and

February 19 in 2018 and 2019, respectively. In sorghum only plots, sorghum was planted on each row at an interplant spacing of 25 cm. In intercropping plots with maize, maize was planted on each row at an interplant spacing of 50 cm whereas sorghum was planted between maize plants to achieve an overall interplant spacing of 25 cm. In intercropping plots with pigeon pea, pigeon pea was planted on every other row at an interplant spacing of 100 cm whereas sorghum was planted on the remaining four rows at an interplant spacing of 25 cm. The three crops were planted by hand using two or three seeds placed 1-1.5 cm deep at each predetermined location and seedlings were thinned to obtain one plant per location 2 weeks after planting. This planting technique was consistent with traditional techniques of Haitian farmers (Levesque 2014). Maize was harvested at maturity and the stalks were removed by hand 10 weeks after planting

sorghum. Pigeon pea plants were allowed to grow until experiment termination. The experimental plots were manually weeded using hoes and furrow-irrigated using irrigation water from the nearby canal as needed. Insecticides were not used to manage insects in these experiments.

Sugarcane Aphid Sampling on Sorghum

Four to 5 weeks after sorghum emergence, the number of sugarcane aphids infesting sorghum plants was recorded approximately every 7 days. On each sampling date, aphids were counted on 10 plants randomly selected on the four center rows in each plot. Three sampling methods were used to estimate sugarcane aphid infestation levels throughout the sorghum growing season because intraplant aphid distribution does not follow a consistent pattern (Elliott et al. 2017). For the first three sampling dates, whole-plant aphid numbers were determined (early season). For the next three sampling dates, aphid numbers were determined on two leaves, one in the lower canopy and one in the upper canopy (middle season). For the remaining three sampling dates, aphid numbers were determined on three leaves, one in the lower canopy, one in the middle canopy, and one in the upper canopy (late season). The first leaf from the base of a plant with >75% its surface green was considered as lower leaf, the newest emerged leaf with a collar was considered as upper leaf, and any leaf located equidistantly from these two leaves was considered as middle leaf. When the flag leaf was present it was considered as the upper leaf (Elliott et al 2017).

Sorghum Growth and Yield Determination

Sorghum plant height and stem diameter were determined on the third sampling date for the early, middle, and late season on 10 plants randomly selected on the four center rows in each plot. Plant height was measured from the plant base to the highest

point of the plant canopy (early season) and to the panicle tip (middle and late season) using a measuring tape. Plant stem diameter was measured 10 cm above the soil surface using a caliper (NEIKO, 0-300mm digital caliper).

Sorghum was harvested approximately 4 weeks following the last sugarcane aphid sampling on January 8, 2019 and June 5, 2019 in the 2018 and 2019 experiments, respectively. Twenty-five plants were randomly selected on the four center rows of each plot and harvested by hand. Each 25-plant sample was weighed using a hanging scale to estimate whole plant fresh biomass. The seeds were manually extracted from each sample and then sun dried during 2 days for 6 hours each day.

Seeds were subsequently weighed using a portable scale (OHAUS, Model CS 200) and moisture was determined using a portable moisture meter (Wile Moisture Wizard 65,

Farmcomp) to estimate plant grain yield adjusted at 13% moisture.

Sugarcane Aphid Natural Enemy Sampling

Plots were sampled for potential sugarcane aphid natural enemies approximately every 7 days for 8 and 5 weeks in 2018 and 2019, respectively, starting at the first sugarcane aphid sampling. On each sampling date, a standard 38-cm diameter sweep net was used to sweep sorghum foliage in each plot. Each sample was composed of thirty sweeps collected from the two center rows of the sorghum only and sorghum- maize plots, and from the fourth and sixth rows in the sorghum-pigeon pea plots. Adults of known aphid predators and parasitoids were identified to family and counted. Sweep net sampling was complemented by direct observations. Sugarcane aphid infestations were observed on 10 randomly selected plants in each plot for 2 to 3 minutes per plant.

Predators observed feeding on sugarcane aphids were sight-identified to family in the field. Aphid mummies were collected and immediately brought to the laboratory at the

American University of the Caribbean, Les Cayes. These mummies were maintained on sorghum leaves in Petri dishes at 25°C and 12:12 (L:D) until parasitoid adult emergence to identify parasitic wasps interacting with sugarcane aphids in the field. Sweep net sampling and direct observations were conducted at different times of the day (morning, mid-day, or late afternoon) among sampling dates so that each plot was sampled at different times to account for potential variation in arthropod diel activity. All adult specimens were preserved in 95% ethanol for future identification to genus or species and subsequent deposition of voucher specimens in the Florida State Collection of

Arthropods (Florida Department of Agriculture and Consumer Services-Division of Plant

Industry, Gainesville, FL).

Sugarcane Aphid Alternate Host Sampling

Plants of all grass species present in the experimental plots and within a 50-m- wide border area surrounding the experimental area were inspected for 1 to 3 hours once a week to identify potential sugarcane aphid alternate hosts. The number of sugarcane aphids found on these potential alternate hosts were recorded. The most common grasses found in the experimental plots and surrounding area were collected and identified.

In addition to the field observations, five itchgrass [Rottboellia cochinchinensis

(Lour.) W.D. Clayton] plants approximately 25 cm tall were collected in the experimental plots with soil surrounding roots and transplanted individually in plastic pots (10 cm diameter x 20 cm height). The plants were maintained in an open field and watered as needed. Five days after transplanting, and after ensuring the plants were free of aphids, each plant was infested with five apterous adult sugarcane aphids collected on sorghum plants using a small paintbrush. Plants were observed twice a day at a 4-hour interval

for 3 days to determine the number of aphids infesting each plant. This experiment was conducted twice, with aphid infestations initiated on March 27 and April 9, 2019.

Statistical Analysis

Sugarcane aphid counts recorded on a per plant or per leaf basis were averaged to obtain the average number of aphids per plant for each plot (early season) or the average number of aphids per leaf for each plot (middle and late season) on each sampling date in the 2018 and 2019 experiments. These average numbers of aphids were used to calculate cumulative aphid days (CAD) for each plot and season using the

3 equation ∑푖=2[(푥푖−1 + 푥푖)/2] × 푡, where x is the average number of aphids per plot on sampling date i, xi-1 is the average number of aphids per plot on the previous sample date, and t is the number of days between sampling dates i-1 and i (Ruppel 1983).

Cumulative aphid days for early, middle, and late season in 2018 and 2019 as affected by treatment were compared using linear mixed models (PROC GLIMMIX, SAS Institute

Inc. 2016). Seasonal assessments and experiments were analyzed separately, thus treatment was a fixed effect whereas block was a random effect for each model.

Plant heights and stem diameters recorded on a per plant basis were averaged to obtain the average height and diameter for each plot and season. Height and diameter as affected by treatment and season were compared for 2018 and 2019 experiments separately using linear mixed models (PROC GLIMMIX, SAS Institute Inc.

2016). Treatment, season, and their two-way interaction were fixed effects whereas block and treatment × block were random effects. Sorghum yield (plant fresh biomass and grain yield) on a per plot basis as affected by treatment in 2018 and 2019 was also compared using linear mixed models. Treatment was a fixed effect whereas block was a

random effect. The number of coccinellids and chrysopids per sweep net sample recorded on a per plot basis for each sampling date was averaged over all sampling dates for each experiment to obtain the average number of coccinellids and chrysopids for each plot. The same model as for sorghum yield was used to compare the number of coccinellids and chrysopids as affected by intercropping. In all models, the Kenward-

Roger adjustment was used to compute error degrees of freedom. The Tukey adjustment at the 5% level was used to interpret mean pairwise comparisons when fixed effects were detected (P < 0.05) (PROC GLIMMIX, SAS Institute Inc. 2016).

Results

Sugarcane Aphid Infestations on Sorghum

In 2018, early-season CAD differed among treatments (P < 0.05), being 4.5-fold lower in sorghum-maize than in sorghum only and sorghum-pigeon pea (Table 2-1).

Differences were detected among treatments for middle-season CAD (P < 0.05), with sorghum-maize exhibiting the lowest CAD that were 5.0-fold lower than in sorghum- pigeon pea (Table 2-1). Late-season CAD also exhibited differences among treatments

(P < 0.05), with sorghum-maize showing 2.6- and 2.9-fold lower CAD than sorghum only and sorghum-pigeon pea, respectively (Table 2-1).

In 2019, early-season CAD differed among treatments (P < 0.05) with sorghum- maize sustaining 28- and 30-fold lower CAD than sorghum only and sorghum-pigeon pea, respectively (Table 2-2). Differences in middle-season CAD were also detected among treatments (P < 0.05, Table 2-2). However, a difference was detected only between sorghum-maize and sorghum-pigeon pea (table 2-2). Differences in late- season CAD were not detected among treatments (P > 0.05, Table 2-2). Differences

between sorghum only and sorghum-pigeon pea were not detected across the three seasons in 2018 and 2019 (Table 2-1 and Table 2-2).

Sorghum Growth and Yield

Sorghum plant height and stem diameter were affected by treatment and season

(P < 0.05, Table 2-3). Plant height across seasons in 2018 and 2019 was 1.4- and 1.8- fold lower, respectively, in sorghum-maize than in sorghum and sorghum-pigeon pea.

Stem diameter was also the smallest in sorghum-maize in 2018 and 2019, with 1.4-fold and 1.9-fold differences, respectively, relative to sorghum and sorghum-pigeon pea

(Table 2-3). However, treatment by season interactions were detected in 2018 and 2019

(P < 0.05, Fig. 2-1, Fig. 2-2). Sorghum plant height was comparable among treatments in early season in both years (Fig. 2-1 A, Fig. 2-2 A); however, sorghum plants in sorghum-maize were shorter than in sorghum only and sorghum-pigeon pea in middle and late season (Fig. 2-1 A, Fig. 2-2 A). Stem diameter in sorghum only and sorghum- pigeon pea was greater than in sorghum-maize in early season in both years. Although this difference persisted, it occurred to a lesser extent in middle season and was detected for late season only in 2019 (Fig. 2-1 B, Fig. 2-2 B).

Differences in sorghum fresh biomass and grain yield were detected (P < 0.05,

Table 2-4) in 2018, with sorghum-maize exhibiting lower fresh biomass and grain yield than sorghum only and sorghum-pigeon pea (Table 2-4). In 2019, differences in sorghum fresh biomass and grain yield were also detected (P < 0.05, Table 2-4).

However, differences in grain yield between sorghum only and sorghum-pigeon pea or sorghum-maize could not be detected although sorghum-maize exhibited the lowest grain yield (Table 2-4). Fresh biomass yield differed among the three treatments, with sorghum-maize being associated with the lowest fresh biomass (Table 2-4).

Sugarcane Aphid Natural Enemies

Coccinellids, chrysopids, syrphids, and braconids were observed. Coccinellids, including H. convergens, and chrysopids were the most commonly collected aphid predators using the sweep net (Table 2-5). The number of coccinellids was lower (P <

0.05) in sorghum-maize than in sorghum-pigeon pea and sorghum only in 2018 and

2019 whereas the number of chrysopids was comparable across intercropping treatments. The number of L. testaceipes adults collected with the sweep net in 2018 was very low with one, zero, and one specimen, in sorghum only, sorghum-maize, and sorghum-pigeon pea, respectively. In 2019, the number of collected L. testaceipes was seven, two, and twelve specimens, in sorghum, sorghum-maize, and sorghum-pigeon pea, respectively. One, four, and eight syrphids were collected in sorghum only, sorghum-maize and sorghum-pigeon pea, respectively, in 2018 whereas, one, zero, and two syrphids were collected in sorghum, sorghum-maize and sorghum-pigeon pea, respectively, in 2019. Direct observations confirmed sweep net sampling with coccinellid, chrysopid, and syrphid larvae observed feeding on aphids. Adults were also observed resting on plant surfaces. Lysiphlebus testaceipes and its pteromalid hyperparasitoid emerged from aphid mummies reared in the laboratory.

Sugarcane Aphid Alternate Hosts

Pangola grass (Digitaria decumbens Stent), Panama crowngrass (Paspalum fimbriatum Kunt), smooth crabgrass [Digitaria ischaemum (Schreb.) Schreb. ex Muhl.], itchgrass [Rottboellia cochinchinensis (Lour.) W.D. Clayton], browntop signalgrass

[Urochloa fusca (Sw.) B.F. Hansen & Wunderlin], and jungle rice [Echinochloa colona

(L.) Link] were found in the experimental plots and surrounding area. Sugarcane aphids were observed on itchgrass throughout the sorghum growing season in the 2018 and

2019 experiments (Table 2-6). When infested, one to four sugarcane aphids were found on individual itchgrass plants (Table 2-6). Although sugarcane aphids were observed on itchgrass in the field, the presence of sugarcane aphids on the potted itchgrass plants was not detected since the first observation following artificial infestation.

Discussion

This study is the first to address aspects of integrated pest management complementing the current approach based on host plant resistance to combat the sugarcane aphid in sorghum in Haiti. Intercropping is a common sorghum production practice in Haitian farming that has assisted in managing insect pests in other agroecosystems (Khan 2006, Agboka et al. 2006, Leclerc et al. 2013, Eckert et al.

2017). The results of this study showed that intercropping sorghum with maize reduces sugarcane aphid infestations on sorghum. Maize plants were 4 weeks old when sorghum was planted. Thus, taller maize plants interspersed with sorghum plants during sorghum development may have acted as a barrier to sugarcane aphid colonization.

Sugarcane aphids have not been observed colonizing maize in Haiti (W. Calvin, personal observation), and aphids landing on maize plants may have left the sorghum- maize plots after initial probing determined maize was not a suitable host plant (Helden and Tjallingii 1993, Klingler et al. 1998, Lazzari et al. 2009, Fartek et al. 2012).

Early planted maize competed with sorghum plants, which negatively affected sorghum growth even after maize harvest and stalk removal. Although sorghum plants in sorghum-maize sustained the lowest sugarcane aphid infestations in this study, yields were the lowest. In contrast, sorghum plants in sorghum-pigeon pea were associated with the highest sugarcane aphid infestations and highest yield. Thus, interplant

competition had considerable effects on yield, which is consistent with observations of grain sorghum intercropped with wild cane (Saccharum spontaneum L.) and Kansas

Orange, a forage sorghum variety (Vesecky et al. 1973). Maize is more competitive than sorghum in terms of water use efficiency (Amanulla 2015). In addition, plant competition is more severe when two grasses are intercropped than when grasses are intercropped with legumes in that similar root systems intensify interplant competition (Dubbs 1971,

Rubio et al. 2001). The fact that maize was planted 4 weeks before sorghum allowed the early establishment of the maize root system, increasing competition with sorghum for water and nutrients. In addition, maize canopy blocked sunlight, which impaired photosynthesis, an important process for plant growth and development (Jagtap et al.

1998). Thus, sorghum plants in sorghum-maize plots were less vigorous than those in sorghum only and sorghum-pigeon pea plots. This lack of vigor might have reduced aphid density, consistent with observations that sorghum planted at a high density sustains relatively lower sugarcane aphid infestations (Singh et al. 2004, Lipsey et al.

2017). In addition, sorghum growing under nitrogen-limited conditions is less suitable for sugarcane aphid development, decreasing fecundity and lifespan (Lama et al. 2019).

This study is the first to observe and identify natural enemies of the sugarcane aphid infesting sorghum in Haiti. Lysiphlebus testaceipes and associated aphid mummies were observed and collected in 2018 and 2019. Chrysopids were common, and syrphid and coccinellid larvae were actively feeding on the aphids. In addition,

Knight (1944) reported L. testaceipes, Cycloneda sanguinea, Scymnus sp., and Baccha clavata F. attacking cotton aphid on cotton and palay rubbervine. These observations suggest that these natural enemies assist in decreasing infestations as seen in previous

studies in North America (Colares et al. 2015a, Rodriguez-del-Bosque et al. 2018,

Delgado-Ramírez et al. 2019). However, a L. testaceipes hyperparasitoid was also present. This may reduce the efficacy of L. testaceipes parasitism against the sugarcane aphid (Ganyo et al. 2012).

Architecturally complex plants generally provide a diversity of food sources and abundant shelter that attract more insects in terms of number of species and individuals than architecturally simple plants (Bianchi et al. 2006, Rundlof and Smith 2006, Speight et al. 2007). Pigeon pea is a broad-leaf shrub with a more complex structure than grasses such as sorghum and maize. Thus, sorghum-pigeon pea intercropping could have been expected to increase sugarcane aphid natural enemy abundance compared to sorghum monoculture or sorghum-maize intercropping. However, coccinellids were more abundant in sorghum monoculture and sorghum-pigeon pea plots than in sorghum-maize plots. The greater number of coccinellids observed in these plots might therefore have only been associated with the numerical response of coccinellids to greater sugarcane aphid densities.

The sugarcane aphid was not observed colonizing grasses other than sorghum in this study. However, the insect was found at low densities on itchgrass (< 5 aphids per infested plant). The corn leaf aphid [Rhopalosiphum maidis (Fitch)], which also infests sorghum, was observed on itchgrass in Honduras (Evans and Halbert 2007), but the presence of sugarcane aphids on itchgrass, to the best of our knowledge, has not been reported previously. In addition, preliminary observations suggest that sugarcane aphids do not survive on itchgrass. Thus, itchgrass, should not be considered as a sugarcane aphid host in Haiti. Consistent with observations in North America (Long et

al. 2018, Nibouche et al. 2018), the sugarcane aphid has been observed colonizing johnsongrass in the vicinity of sorghum fields in western Haiti (M. D. Dorval, Personal communication). Johnsongrass can be a suitable alternate host for sugarcane aphid in

Haiti since this aphid can develop comparatively well on johnsongrass (de Souza and

Davis 2019). Johnsongrass was not present in experimental plots of this study.

In conclusion, this study suggests that intercropping sorghum with maize reduces sugarcane aphid infestations on sorghum. However, competition among maize and sorghum plants is a concern because it negatively affects sorghum yield. A more efficient sorghum-maize intercropping design may decrease sugarcane aphid infestations and improve yield. The results of this study also provide evidence that natural enemies of the sugarcane aphid are present in Haiti. However, natural enemy abundance and the level of biological control they may provide was not determined in this study. Although alternate hosts of the sugarcane aphid other than johnsongrass may be present in Haiti, our study showed low evidence with respect to common grasses. Future research should investigate different intercropping designs with maize.

In addition, future studies aiming to determine the population dynamics, the abundance and the effects of beneficial insects on the pest must be considered. This will provide further information for the development of an effective integrated management program for sugarcane aphid in Haiti.

Table 2-1. Cumulative aphid days (CAD) for sugarcane aphids infesting sorghum in sorghum, sorghum-maize, and sorghum-pigeon pea plots, Jonc-Labeille, Haiti, October-December 2018. CAD Treatment Early season* Middle season* Late season* [LS means ± 69.5 [LS means ± 36.8 [LS means ± 89.7 (SE)] (SE)] (SE)] Sorghum 616.4a 145.4ab 398.8a Sorghum-maize 136.0b 45.6b 150.9b Sorghum-pigeon 612.9a 226.4a 439.7a pea F 15.8 7.6 6.1 df 2, 15 2, 10 2, 10 P > F <0.001 0.01 0.019 *Means with the same letter in a column are not significantly different (Tukey adjustment, α = 0.05).

Figure 2-1. Sorghum plant height (A) and stem diameter (B) (LS means) as affected by intercropping treatment, Jonc-Labeille, Haiti, 2018.

Table 2-2. Cumulative aphid days (CAD) for sugarcane aphids infesting sorghum in sorghum, sorghum-maize, and sorghum-pigeon pea plots, Jonc-Labeille, Haiti, March-May 2019. CAD Treatment Early season* Middle season* Late season* [LS means ± 120.4 [LS means ± 27.0 [LS means ± 41.2 (SE)] (SE)] (SE)] Sorghum 983.1a 49.6ab 205.8a Sorghum-maize 35.3b 10.6b 116.1a Sorghum-pigeon 1048.6a 118.a 148.6a pea F 22.2 4.1 1.2 df 2, 15 2, 15 2, 15 P > F <0.001 0.039 0.324 *Means with the same letter in a column are not significantly different (Tukey adjustment, α = 0.05).

Figure 2-2. Sorghum plant height (A) and stem diameter (B) (LS means) as affected by intercropping treatment, Jonc-Labeille, Haiti, 2019.

Table 2-3. Sorghum plant height and stem diameter comparisons in intercropping experiments, Jonc-Labeille, Haiti, 2018 and 2019. 2018 2019 Plant height* Stem diameter* Plant height* Stem diameter* Treatment (cm) [LS (mm) [LS (cm) [LS (mm) [LS means ± 4.1 means ± 0.8 means ± 4.5 means ± 0.4 (SE)] (SE)] (SE)] (SE)] Sorghum 149.5a 14.8a 155.7a 14.6b Sorghum- 107.7b 10.2b 84.5b 8.1c maize Sorghum- 147.4a 14.7a 156.7a 16.2a pigeon pea F 33.5 12.0 84.0 115.0 df 2, 15 2, 15 2, 15 2, 15 P > F <0.001 <0.001 <0.001 <0.001 Season [LS means ± [LS means ± 0.7 [LS means ± [LS means ± 0.3 2.9 (SE)] (SE)] 3.1 (SE)] (SE)] Early 65.2c 11.2b 60.8c 10.5b Middle 149.4b 13.9a 149.9b 14.1a Late 190.0a 14.6a 186.2a 14.3a F 900.1 6.1 902.7 134.4 df 2, 30 2, 30 2, 30 2, 30 P > F <0.001 0.006 <0.001 <0.001 * Means with the same letter in a column are not significantly different (Tukey adjustment, α = 0.05).

Table 2-4. Sorghum yield comparisons in intercropping experiments, Jonc-Labeille, Haiti, 2018 and 2019. 2018 2019 Grain yield* Fresh biomass* Grain yield* Fresh biomass* Treatment (g/plant) (kg/plant) (g/plant) (kg/plant) [LS means ± [LS means ± [LS means ± [LS means ± 7.6 (SE)] 0.02 (SE)] 2.7 (SE)] 0.02 (SE)] Sorghum 16.6a 0.4a 15.3ab 0.4b Sorghum- 6.4b 0.2b 6.1b 0.2c maize Sorghum- 14.0a 0.4a 24.7a 0.5a pigeon pea F 20.5 5.6 12.0 50.6 df 2, 10 2, 10 2, 15 2, 15 P > F <0.001 < 0.001 <0.001 < 0.001 * Means with the same letter in a column are not significantly different (Tukey adjustment, α = 0.05).

Table 2-5. Mean number of insects collected with a sweep net in intercropping experiments, Jonc-Labeille, Haiti, 2018 and 2019. 2018 2019 Chrysopidae* Coccinellidae* Chrysopidae* Coccinellidae* Treatment (No./plot) (No./plot) (No./plot) (No./plot) [LS means ± [LS means ± [LS means ± [LS means ± 0.3 (SE)] 0.19 (SE)] 0.13 (SE)] 0.19 (SE)] Sorghum 1.15a 2.0a 0.6a 0.8ab Sorghum-maize 1.3a 1.1b 0.5a 0.2b Sorghum- 1.5a 2.4a 0.6a 1.3a pigeon pea F 0.3 14.0 0.2 8.2 df 2, 15 2, 10 2, 10 2, 10 P > F 0.756 0.001 0.852 <0.001 * Means with the same letter in a column are not significantly different (Tukey adjustment, α = 0.05).

Table 2-6. Number of sugarcane aphids found on itchgrass, Jonc-Labeille, Haiti, 2018 and 2019. Experiment Date Number of infested Number of aphids per plants infested plant 2018 09/13/18 2 2 3 09/21/18 2 1 1 09/26/18 1 1 10/03/18 1 3 10/18/18 0 -- 10/24/18 0 -- 11/07/18 1 1 11/14/18 1 2 11/29/18 1 0 2019 03/06/19 4 1 2 1 1 03/15/19 2 1 1 03/21/19 3 4 3 1 03/27/19 3 4 2 1 04/05/19 0 -- 04/11/19 0 -- 04/16/19 4 1 1 1 1 04/23/19 0 -- 05/01/19 1 1

CHAPTER 3 EFFECTS OF BIOLOGICAL INSECTICIDES ON SUGARCANE APHID (HEMIPTERA: APHIDIDAE) INFESTATIONS ON SORGHUM

Sugarcane aphid [Melanaphis sacchari (Zehntner)] management on sorghum

[Sorghum bicolor (L.) Moench] in the United States has primarily relied on the use of resistant varieties and application of conventional insecticides (Villanueva et al. 2014,

Brown et al. 2015, Knutson et al. 2015, Brewer et al. 2016). However, excessive use of these insecticides targeting the nicotinic acetylcholine receptor to manage infestations can induce the development of insecticide resistant aphid populations (Bowling et al.

2016, Szczepaniec 2018, Etheridge et al. 2019, IRAC 2019). In addition, conventional insecticides are persistent and their accumulation in nature can give rise to important environmental concerns (Edwards and Adams 1970, Wang et al. 2011, Waskom et al.

2017). In contrast, biological insecticides have a diversity of modes of action allowing rotation to mitigate the development of insecticide resistance, and they do not accumulate in the environment (Copping and Menn 2000, Chandler et al. 2011). In addition, the adoption of biological insecticides may offer reduced-risk production practices in developing countries where farmers are not well equipped and trained to use pesticides (USAID 2010). Thus, biological insecticides may be a suitable tactic to be integrated into a pest management program in the United States and in developing countries such as Haiti.

Biological insecticides based on botanical extracts, entomopathogenic fungi, or entomopathogenic bacteria and their toxins have adverse effects against insect pests including aphids (Stark and Walter 1995, Buss and Park-Brown 2009, Selvaraj and

Kaushik 2014, Kuhar and Doughty 2016, Yadav et al. 2016). For instance, insecticides based on neem [Azadirachta indica (A. Juss.)] seed extracts can be effective aphicides

(Lowery and Isman 1994, Krais and Cullen 2008, Yadav et al. 2016). Neem seed oil and its main insecticidal component, azadirachtin, cause nymphal mortality, prolong developmental time, and reduce fecundity of the soybean aphid [Aphis glycines

(Matsumura)] (Kraiss and Cullen 2008). A mixture of neem seed oil and azadirachtin reduces the life span and fecundity of the pea aphid [Acyrthosiphon pisum (Harris)] and can be lethal by affecting the molting process (Stark and Walter 1995).

Pyrethrins, a mixture of six active compounds extracted from Chrysanthemum cinerariifolium (Trevir.) Vis. plants, have provided control of insect pests including aphids (Casida 1980, Buss and Park-Brown 2009, Khater 2012, Singh 2014). Field experiments showed that pyrethrins can control the pea aphid and alfalfa plant bug

[Adelphocoris lineolatus (Goeze)] in Bulgaria (Niklova 2016). In addition, pyrethrins negatively affect the brown marmorated stink bug [Halyomorpha halys (Stål)] in laboratory bioassays (Lee et al. 2014, Morehead and Kuhar 2017).

Oil extracted from vetiver grass [Chrysopogon zizanioides (L.) Roberty] is composed of -vetivones, -vetivones, khusinol, khusilal, diethyl phthalate, vetiselinol, khusimol, isovelencenol, and vetivenic acid. This botanical extract has been reported as an effective repellent and toxicant of insect pests including the red imported fire ant

(Solenopsis invicta Buren), German cockroach [Blattella germanica (L.)], blacklegged tick [Ixodes Scapularis (Say)], and Formosan subterranean termite [Coptotermes formosanus (Shiraki)] (Zhu et al. 2003, Henderson et al. 2005a, 2005b). In addition, vetiver grass extracts have caused more than 50% mortality in the cowpea weevil

[Callosobruchus maculattus (F.)] under laboratory conditions (Pangnakorn 2009).

Beauveria bassiana (Bals. -Criv.) Vuill. is an entomopathogenic fungus that was found naturally in sugarcane aphids in several regions in Mexico (Zambrano-Gutierrez et al. 2019). Beauveria bassiana CKB-48 has caused more than 90% nymphal mortality in the sugarcane aphid, corn leaf aphid (Rhopalosiphum maidis Fitch), and five other aphid pest species in laboratory experiments (Maketon et al. 2013). In another laboratory study, B. bassiana negatively affected the greenbug [Schizaphis graminum

(Rondani)], bird cherry-oat aphid [Rhopalosiphum padi (L.)], cabbage aphid

[Brevicoryne brassicae (L.)], and mustard aphid [Lipaphis erysimi (Kaltenbach)] (Akmal et al. 2013). In addition, greenhouse and field experiments showed that B. bassiana strain HaBa can effectively control the cowpea aphid [Aphis craccivora (Das)] (Selvaraj and Kaushik 2014). Another fungus, Isaria fumosorosea (Wize) Brown & Smith provides as much as 100% control of the brown citrus aphid [Toxoptera citricidus (Kirkaldy)]

(Hunter et al. 2011). Isaria fumosorosea has also shown insecticidal effects against the red palm weevil [Rhynocophorus ferrugineus (Olivier)] by causing as much as 100% egg and larval mortality (Sabbour and Abdel-Raheem 2014). Gandarilla-Pacheco et al.

(2015) reported that I. fumosorosea can cause 40 and 57% mortality in neonate beet armyworm [Spodoptera exigua (Hübner)] and corn earworm [Helicoverpa zea (Boddie)], respectively.

The bacteria Chromobacterium subtsugae strain PRAA4-1T can inhibit feeding or cause mortality in the Colorado potato beetle [Leptinotarsa decemlineata (Say)], the sweet potato whitefly [Bemisia tabaci (Gennadius)], and seven other insect pest species in laboratory bioassays (Martin et al. 2007). Chromobacterium subtsugae also has adverse effects on the melon aphid (Aphis gossypii Glover) (Kuhar and Doughty 2016)

and the black pecan aphid [Melanocallis caryaefoliae (Davis)] (Shapiro-Ilan et al. 2013).

Other bacteria, Burkholderia spp. strain A396, showed suitable control of the cranberry fruitworm [Acrobasis vaccinia (Riley)] in blueberry (Vaccinium spp.) (Wise et al. 2015).

In addition, laboratory bioassays showed that Burkholderia spp. strain A396 can cause up to 85% beet armyworm mortality (Cordova-Kreylos et al. 2013).

Azadirachtin, pyrethrins, B. bassiana, I. fumosorosea, C. subtsugae, and

Burkholderia spp. are commercially formulated as biological insecticides in the United

States. In addition, Haiti is the largest producer of vetiver in the world (Belhassen et al.

2015). Thus, commercial biological insecticides and vetiver oil may represent an additional tactic to consider in an integrated pest management strategy for the sugarcane aphid in sorghum in the United States and the Caribbean, including Haiti. In this study, the efficacy of biological insecticides registered on numerous crops in the

United States, as well as vetiver oil, was evaluated in the laboratory, greenhouse, and field for control of the sugarcane aphid.

Materials and Methods

Experimental Treatments

All experimental treatments were mixed in deionized water 30-60 minutes before use. Concentrations for the commercial biological insecticides were determined to be consistent with the highest registered field rates applied at a volume of application of

187 l/ha. Botanical treatments included azadirachtin (Molt-X® EC, BioWorks, Victor, NY) at an equivalent of 24.5 g (AI)/ha, pyrethrins (Pyganic® 5.0 EC, Valent U.S.A, Walnut

Creek, CA) at an equivalent of 56.0 g (AI)/ha, and Haitian vetiver essential oil

(Floracopeia, Grass Valley, CA) at 2% v/v. The vetiver oil solution was warmed at 38°C for 30 minutes and vigorously shaken immediately before use to mix the oil in water.

Fungal treatments included, Beauveria bassiana strain GHA (BotaniGard® ES,

BioWorks) at an equivalent of 236.2 g (AI)/ha and Isaria fumosorosea Apopka strain 97

(PFR-97™ 20% WDG, Certis USA, Columbia, MD) at an equivalent of 448.3 g (AI)/ha.

Bacterial treatments included Chromobacterium subtsugae strain PRAA4-1T

(Grandevo®, Marrone Bio Innovations, Davis, CA) at an equivalent of 1008.8 g (AI)/ha and Burkholderia spp. strain A396 (Venerate®, Marrone Bio Innovations) at an equivalent of 8834.7 g (AI)/ha. In addition, a commercial pre-mix of Beauveria bassiana strain GHA and pyrethrins (BotaniGard® Maxx, BioWorks) at 1.12 + 15.4 g (AI)/ha was included. The butenolide flupyradifurone (Sivanto™ prime, Bayer CropScience,

Research Triangle Park, NC) at an equivalent of 102.4 g (AI)/ha served as conventional insecticide standard.

Aphid Colony

A sugarcane aphid colony was initiated using a single apterous aphid collected on May 28, 2018 from a field of sweet sorghum (M-81E variety, Broadhead et al. 1981) at the University of Florida Institute of Food and Agricultural Sciences Everglades

Research and Education Center (UF/IFAS EREC) in Belle Glade, FL. The aphids were reared on 2-4-week-old M-81E sorghum plants maintained in plastic pots (20.3-cm top diameter, 14.2-cm-deep) filled with potting soil (Miracle-Gro All Purpose Potting Mix,

Scotts Miracle-Gro Company, Marysville, OH) at a density of one plant per pot. The plants were kept in a popup rearing cage (61 x 61 x 142 cm, Bioquip, Rancho

Dominguez, CA) in a greenhouse. Once a week, eight plants were infested with 15 adults using a fine paintbrush. As needed, as many as 15 additional sorghum plants were infested with 15 adults per plant 1 day before laboratory and greenhouse

experiments to produce 1-day-old nymphs for infestation in those experiments. Each adult produced an average of four nymphs in 1 day.

Laboratory Experiment

An experiment was conducted during summer 2018 at the UF/IFAS EREC to evaluate the effects of the seven biological insecticides, vetiver oil, and flupyradifurone on sugarcane aphid mortality under laboratory conditions. Sorghum leaves collected from the upper canopy of plants in a 12-week-old field of M-81E were used in the laboratory to prepare leaf discs 7.5 cm in diameter. Although the field sustained sugarcane aphid infestations between 1 and 4 weeks after planting, plants were free of aphids at the time of leaf collection.

The nine treatments, as well as a non-treated control consisting of deionized water, were applied to individual leaf discs using a 60-ml amber glass bottle with mist sprayer (Katzco, Monroe, NY). Each side of a leaf disc received two sprays. The sprayer delivered 0.15 ml of solution for each spray on average. Thus, each leaf disc received 0.60 ml of solution, a volume equivalent to that of a plant being treated with

187 l/ha of spray solution in a field with 76,850 plants/ha. Each leaf disc, which contained the midrib, was placed abaxial surface up in a 9-cm plastic Petri dish on top of a layer of filter paper moist with deionized water. Solutions were allowed to dry before aphid infestation.

Five 1-day-old nymphs from the sugarcane aphid colony were placed at the center of each leaf disc using a fine paintbrush. Aphid mortality was determined 6, 24,

48, and 72 hours following infestation. An aphid showing no perceptible movement after being prodded for 2-3 seconds with a fine paintbrush was considered dead. Three bioassays were conducted, each with a different aphid cohort. Each bioassay included

2-4 replicates, for a total of 10 replicates. Each replicate consisted of 10 Petri dishes, one for each treatment and the non-treated control, placed on an individual wire shelf in an environmentally-controlled room at 26°C, 40-60% RH, 12:12 (L:D).

Greenhouse Experiments

Two experiments were conducted during summer 2018 at the UF/IFAS EREC to evaluate the effects of the seven biological insecticides, vetiver oil, and flupyradifurone on sugarcane aphid infestations developing on sorghum plants under greenhouse conditions. In the first experiment, the effects of treatments applied to sorghum plants already infested with aphids were evaluated whereas the effects of treatments applied to sorghum plants prior to aphid infestation were evaluated in a second experiment. In the two experiments, potted 2-week-old sorghum plants were used (see Aphid Colony section) and each plant was infested with five 1-day-old nymphs on the topmost leaf with a visible collar using a fine paintbrush. The nine treatments, as well as a non- treated control consisting of deionized water, were applied to individual plants using mist sprayers consistent with the laboratory experiment. Each plant was sprayed on two opposite sides, upward and downward, receiving 0.60 ml of solution in total. Each experiment included five assays, each conducted using a different cohort for sorghum plants and aphids.

In each assay of the first experiment, each treatment or the non-treated control was sprayed on three plants infested with aphids 15-30 minutes earlier. The three plants for each treatment were immediately placed together in a screened tent-like cage

(51 x 51 x 51 cm, Bug Dorm 2, Bioquip, Rancho Dominguez, CA). Thus, there were 10 cages with three plants in each that were placed in a random order on a bench in the greenhouse for each assay. In each assay of the second experiment, a comparable

method was used; however, sprayed plants were allowed to dry for 15-30 minutes before being infested with aphids and placed in cages. After 7 days, all plants in an assay were removed from the cages for whole-plant aphid counts.

Field Experiments

Three experiments were conducted at the UF/IFAS EREC during spring 2018, fall 2018, and spring 2019 to further evaluate the effects of the seven biological insecticides and flupyradifurone on sugarcane aphid infestations in the field. Eight treatments (spring and fall 2018) and nine treatments (spring 2019), including a non- sprayed control, were evaluated in a randomized complete block design with four blocks

(1 replication per block). Treatments were assigned to plots four rows wide (76.2-cm row spacing) and 10 m long in a field of sorghum variety M-81E planted at density of

76,850 seeds/ha.

Insecticide treatments were applied using a CO2-pressurized backpack sprayer with a two-row boom equipped with four TeeJet XR 8002VS nozzles (TeeJet

Technologies, Wheaton, IL) spaced 38 cm apart and calibrated to deliver 187 l/ha at

207 kPa. Treatments were initiated at first sign of sugarcane aphid infestations when plants were at the V3, early boot, and V8 stages in spring 2018, fall 2018, and spring

2019, respectively. In the spring 2018 experiment, insecticides were applied twice, 5 days apart, whereas in the two subsequent experiments insecticides were applied four times, 2-5 days apart over 2 weeks.

Aphid counts were taken from ten plants randomly selected on the two center rows of each plot in each experiment. In the spring 2018 experiment, whole-plant aphid numbers were determined. In addition to a pretreatment count 1 day before the first insecticide application, one aphid count was taken 2 days after the last application. The

experiment was subsequently terminated because excessive rainfall prevented additional insecticide applications and the aphid population declined considerably. In the fall 2018 and spring 2019 experiments, aphid numbers were determined for each plant on two leaves, one in the lower canopy and one in the upper canopy. The first leaf from the base of a plant with >75% its surface green was considered as the lower leaf and the newest emerged leaf with a collar or the flag leaf, if present, was considered as the upper leaf. In addition to a pretreatment count 1 day before the first insecticide application, three weekly aphid counts were taken starting on the day of the second insecticide application or 2 days after for the fall 2018 and spring 2019 experiments, respectively.

Statistical Analysis

Data from all experiments were analyzed using linear mixed models (PROC

GLIMMIX, SAS Institute Inc. 2016). For the laboratory experiment, aphid mortality expressed as the percentage of dead aphids was compared with a model including insecticide treatment, observation time, and their two-way interaction as fixed effects and bioassay, replicate(bioassay), and treatment × replicate(bioassay) as random effects. For each greenhouse experiment, whole-plant aphid numbers were compared with a model including insecticide treatment as a fixed effect and assay and treatment × assay as random effects. For the spring 2018 field experiment, whole-plant aphid numbers averaged on a per plot basis were compared with a model including insecticide treatment as a fixed effect and block as a random effect. For the fall 2018 and spring 2019 experiments, aphid numbers per leaf averaged on a per plot basis were compared with a model including observation date and the treatment by

observation date interaction as additional fixed effects and block and treatment × block as additional random effects. The Kenward-Roger adjustment for denominator degrees of freedom was used to correct for inexact F distributions in all models. The Tukey adjustment ( = 0.05) was used to assist in interpreting pairwise differences in least square means. When a two-way interaction was significant at  = 0.05, the SPLICE and

SPLICEDIFF options were used to assist in comparing treatment least square means at each observation time or date (PROC GLIMMIX, SAS Institute Inc. 2016).

Results

Laboratory Experiment

Differences in sugarcane aphid mortality were detected (F = 33.5; df = 9, 80.8; P

< 0.001) among treatments across observation times (Fig. 3-1). The non-treated control had the lowest mortality [12.3 ± 6.7 (SE) %] whereas the highest mortality was observed on flupyradifurone-treated leaf discs [98.7 ± 6.7 (SE) %]. Mortalities for the pre-mix of B. bassiana and pyrethrins [94.2 ± 6.7 (SE) %], pyrethrins [88.3 ± 6.7 (SE) %], and C. subtsugae [79.9 ± 6.7 (SE) %)] were the highest among biological insecticides and were not different from the mortality observed on flupyradifurone-treated leaf discs. Beauveria bassiana, azadirachtin, and vetiver oil were associated with intermediate mortality, with

66.3%, 44.8%, and 41.8%, respectively. Mortalities on leaf discs treated with

Burkholderia spp. and I. fumosorosea were not different (P > 0.05) than mortality on non-treated discs.

Mortality across treatments generally increased between 6 and 72 hours after assay initiation (F = 92.8; df = 3, 249.9; P < 0.001), with mortality in non-treated Petri dishes increasing from 0.4 ± 7.3 (SE) % at 6 hours to 20.0 ± 7.6 (SE) % at 72 hours

(Fig. 3-1). However, a treatment by observation time interaction was detected (F = 9.3; df = 27, 250.6; P < 0.001), indicating that the effects of treatments varied with observation time (Fig. 3-1). Flupyradifurone, pyrethrins, the pre-mix of B. bassiana + pyrethrins, and C. subtsugae showed substantial effects on the aphids starting 6 hours after assay initiation and mortality increased by 5.4-16.8% 72 hours after assay initiation. However, the remaining treatments were associated with relatively low mortality at 6 hours. Mortality subsequently increased at 72 hours by a minimum of

72.0% for B. bassiana and a maximum of 34.2-fold for azadirachtin (Fig. 3-1).

Greenhouse Experiments

The number of sugarcane aphids infesting sorghum plants differed (P < 0.05) among treatments in the two greenhouse experiments (Table 3-1). In the first experiment (aphid-infested plants sprayed), aphid infestation levels decreased by 100% on flupyradifurone-treated plants relative to non-treated plants. Plants treated with azadirachtin, pyrethrins, and the B. bassiana + pyrethrins premix sustained 98.2, 93.4, and 82.3% less sugarcane aphids, respectively, than non-treated plants (Table 3-1). In the second experiment (non-infested plants sprayed), flupyradifurone also decreased infestation by 100%. Azadirachtin and pyrethrins were the only other treatments with measurable effects on sugarcane aphid infestations, with 96.9 and 65.0% less aphids, respectively, than the non-treated plants (Table 3-1). Although the pre-mix of B. bassiana + pyrethrins was different than the non-treated control in the first experiment, differences were not detected (P > 0.05) in the second experiment (Table 3-1).

Field Experiments

Differences in sugarcane aphid infestation levels were detected (P < 0.05) among treatments for the single post-treatment observation in spring 2018 and across

the three post-treatment observations in spring 2019 (Table 3-2). In spring 2018, infestation levels in flupyradifurone-treated plots were 95.0% lower than in azadirachtin- treated plots. In spring 2019, infestation levels in flupyradifurone-treated plots were

99.1% lower than in I. fumosorosea-treated plots. However, aphid infestation levels in non-sprayed plots were not different than those in any treated plots in the two experiments (Table 3-2). In the fall 2108 experiment, although differences were not detected (P > 0.05) among treatments, flupyradifurone-treated plots sustained the lowest sugarcane aphid infestation levels, consistent with the other two experiments

(Table 3-2).

Infestation levels across treatments increased (P < 0.05) between the second and third weekly post-treatment observation dates for the fall 2018 and spring 2019 experiments (Table 3-2). In addition, a treatment by observation date interaction was detected (F = 2.1; df = 16, 54; P = 0.024) for the spring 2019 experiment. Although infestation levels were not different among treatments for the first and second post- treatment observation dates, differences were detected (P < 0.05) for the third date.

However, infestation levels in non-sprayed plots were not different than those in any other treated plots for that date (Fig. 3-2).

Discussion

This study, to the best of our knowledge, is the first to provide a comprehensive assessment of biological compounds that may benefit sorghum farmers in the United

States and Haiti. Since the emergence of the sugarcane aphid as a major sorghum pest in North America, farmers have relied on a limited number of conventional insecticides with similar modes of action to manage the aphid (Bowling et al. 2016, IRAC 2019).

However, biological insecticides may represent an additional management tactic to

combat this pest because they have multiple modes of action (Marrone 2019) and can be effective aphicides (Stark and Walter 1995, Kraiss and Cullen 2008, Maketon et al.

2013, Kuhar and Doughty 2016).

Azadirachtin and pyrethrins negatively affected sugarcane aphid infestations in laboratory and greenhouse experiments in this study. These results are consistent with insecticidal activity observed on numerous insect pests in previous laboratory and field studies (Kraiss and Cullen 2008, Niklova 2016, Morehead and Kuhar 2017, Diaz-Najera et al. 2018). In the laboratory experiment, the pyrethrins caused >80% mortality at 6 hours after assay initiation whereas azadirachtin caused comparable mortality at 72 hours after assay initiation. These observations suggest that azadirachtin does not have an immediate effect on the sugarcane aphid likely because of its repellent and growth regulator activity (Kraiss and Cullen 2008, Buss and Park-Brown 2009). In the greenhouse experiments, mainly adult aphids were observed on azadirachtin-treated plants sustaining low aphid infestation levels (W. Calvin, personal observations), suggesting that azadirachtin might have an effect on the development and reproductive capability of the sugarcane aphid nymphs. Other studies showed that neem seed oil and azadirachtin cause nymphal mortality, prolong developmental time, and reduce fecundity of the soybean aphid (Kraiss and Cullen 2008). In contrast, pyrethrins have immediate effects on exposed insects.

Vetiver oil caused nearly 60% sugarcane aphid mortality in the laboratory experiments at 72 hours although very low sugarcane aphid mortality was associated with vetiver oil after 6 hours. However, the sugarcane aphid infestation levels in vetiver oil-treated sorghum were comparable to the non-treated control in the greenhouse. The

mode of action of vetiver oil against the sugarcane aphid is unknown. However, vetiver oil repellency and toxicity to several arthropod pests have been observed (Zhu et al.

2003, Henderson et al. 2005a, 2005b). Haiti is the first vetiver oil producer worldwide

(Belhassen et al. 2015) and large quantities of byproducts are produced daily. These byproducts may contain some levels of vetiver oil and may serve as a potential low-cost insecticide to smallholders in Haiti. In addition, intercropping vetiver grass with other crops may cause deleterious effects to pest infestations. For instance, the spotted stem borer [Chilo partellus (Swinhoe)] prefers to lay eggs on vetiver grass, which assists in controlling the insect because of a decrease in offspring survival (Van den Berg et al.

2003). Thus, the use of vetiver oil and vetiver grass may play a role in an integrated sugarcane aphid management.

Beauveria bassiana strain GHA negatively affected sugarcane aphids in the laboratory and greenhouse experiments but to a lesser extent than in a previous laboratory study showing that B. bassiana CKB-48 can cause nymphal mortality approaching 100% in the sugarcane aphid (Maketon et al. 2013). The pre-mix of B. bassiana + pyrethrins was among the most effective biological insecticides in the laboratory and greenhouse experiments, and negatively affected sugarcane aphids to a greater extent than did B. bassiana alone despite the reduced rate in the pre-mix.

Similarly, Reddy and Antwi (2016) showed that B. bassiana applied alone was less effective against wheat head armyworm [Dargida diffusa (Walker)] larvae than when mixed with other plant extracts, indicating that B. bassiana is more effective when combined with other insecticides. The other fungus, I. fumosorosea, did not have measurable effects on the sugarcane aphid in any experiment of this study. Although I.

fumosorosea has negative effects on selected insect pests (Hunter et al. 2011), this biological insecticide should not be considered in a sugarcane aphid management program. In contrast, further research on B. bassiana alone or in combination with other biological insecticides is warranted for consideration in a management program.

The bacterial insecticide C. subtsugae showed adverse effects on sugarcane aphid only in the laboratory experiment whereas Burkholderia spp. did not show measurable effects in any of the experiments of this study. It is likely that the activity of

C. subtsugae on the sugarcane aphid is limited to narrow environmental conditions because in a previous field experiment, C. subtsugae was also uneffective against the sugarcane aphid (Studebaker and Jackson 2017). Nonetheless, other studies showed that C. subtsugae can be deleterious to numerous pest species including aphids

(Shapiro-Ilan et al. 2013, Andon and Shetlar 2015, Kuhar and Doughty 2016). To the best of our knowledge, Burkholderia spp. has not been previously tested on any aphid species. However, previous studies showed that Burkholderia spp. can have adverse effects on the cranberry fruitworm and beet armyworm (Cordova-Kreylos et al. 2013,

Wise et al. 2015). These results also suggest that C. subtsugae and Burkholderia spp. should not be considered in a sugarcane aphid management program.

The conventional insecticide, flupyradifurone, used as a standard treatment had consistent efficacy against the sugarcane aphid across the laboratory, greenhouse, and field experiments. This insecticide was consistently associated with the highest mortality or lowest infestation levels although differences with other treatments were not always detected. In contrast, the efficacy of biological insecticides was not consistent across the experiments. Some biological insecticides showed high efficacy in the laboratory

experiment with intermediate efficacy in the greenhouse experiments and little to no efficacy in the field experiments, which is consistent with Edelson et al. (2002). This decrease in biological insecticide efficacy might be partially associated with a difference in spray coverage because the sorghum plants in the field might receive lower coverage than the sorghum leaf discs used in the laboratory experiment and the potted sorghum plants used in the greenhouse experiments (Shapiro-Ilan et al. 2013). In addition, the environmental conditions to which the insecticides were exposed in the greenhouse and the field likely decreased their efficacy. Temperature, precipitation, wind, solar radiation, and microbial activity are factors that influence insecticides efficacy by causing rapid degradation of their active ingredients (Mulla and Su 1999). For instance, pyrethrins might poorly persist in the field because of their photo-instability (Copping and Menn

2000). Pyrethrin1, which is one of the pyrethrins components, breaks down rapidly on plant surfaces when exposed to light with a half-life of 11.8 hours and 12.9 hours in water and on soil surfaces, respectively. Approximately 3% of pyrethrins can remain on plants 5 days after treatment. Azadirachtin is also subject to rapid degradation by light and microbial activity with a half-life of 1 to 2.5 days on plant leaves (NPIC 2012, 2014).

However, a previous study showed that azadirachtin can be active up to 21 days after treatment comparing to 35 days after treatment for cyfluthrin, tebufenozide, and diflubenzuron (Webb et al. 1998). Laboratory experiments showed that azadirachtin and

B. bassiana can have greater insecticidal activity on the differential grasshopper

[Melanoplus differentialis (Thomas)] when applied at temperature between 25°C and

35°C for azadirachtin and between 15°C and 25°C for B. bassiana (Amarasekare and

Edelson 2004). Thus, it can be assumed that environmental conditions in the greenhouse and field were far from optimal for the performance of the insecticides.

This study suggests that biological insecticides including pyrethrins, azadirachtin,

B. bassiana, and the pre-mix of B. bassiana and pyrethrins could potentially control sugarcane aphid infestations on sorghum if they are applied under acceptable environmental conditions with good coverage. Therefore, further studies should address application methods, environmental conditions, and rates conducive to optimal efficacy of these insecticides in the field. The use of these biological insecticides in a sugarcane aphid pest management program would allow reduced-risk sorghum production and insecticide resistance mitigation. The use of these insecticides would also be very important in organic sorghum production.

Figure 3-1. Sugarcane aphid mortality (LS means) over time as affected by the application of seven biological insecticides, vetiver oil, and a conventional insecticide under laboratory conditions, summer 2018, Belle Glade, FL.

Table 3-1. Sugarcane aphid infestation levels on sorghum plants as affected by the application of biological insecticides and vetiver oil in the greenhouse, Belle Glade, FL, summer 2018. In the first experiment, treatments were applied to sorghum plants infested with aphids. In the second experiment, treatments where applied to sorghum plants before infestation with aphids. First experiment Second experiment Treatment No. aphids/plant* No. aphids/plant* [LS means ± 8.0 (SE)] [LS means ± 10.8 (SE)] Non-treated 59.3a 61.7a Azadirachtin 1.1c 1.9c Pyrethrins 3.9c 21.6bc Vetiver oil 48.1ab 58.7a Beauveria bassiana 24.9bc 43.7ab Beauveria bassiana + pyrethrins 10.5c 47.7ab Isaria fumosorosea 45.3ab 34.5abc Chromobacterium subtsugae 46.8ab 54.1ab Burkholderia spp. 51.6ab 51.9ab Flupyradifurone 0.0c 0.0c F 12.9 8.6 df 9, 36 9, 36 P > F <0.001 <0.001 *Means with the same letter in a column are not significantly different (Tukey adjustment, α = 0.05).

Table 3-2. Sugarcane aphid infestation levels as affected by insecticide treatment and post-treatment observation date in three field experiments, Belle Glade, FL, 2018 and 2019. Spring 2018 Fall 2018 Spring 2019 Treatment No. aphids/plant* No. aphids/leaf* No. aphids/leaf* [LS means ± 3.5 [LS means ± 15.7 [LS means ± 4.2 (SE)] (SE)] (SE)] Non-sprayed 14.6ab 38.0a 10.4ab Azadirachtin 20.2a 14.8a 7.7ab Pyrethrins - 6.8a 14.0ab Beauveria bassiana 13.3ab 47.8a 14.2ab Beauveria bassiana + pyrethrins 12.3ab 47.3a 15.1ab Isaria fumosorosea 10.0ab - 25.3a Chromobacterium subtsugae 16.1ab 46.5a 12.6ab Burkholderia spp. 14.4ab 35.0a 8.9ab Flupyradifurone 1.0b 0.5a 0.2b F 2.8 1.7 3.1 df 7, 21 7, 21 8, 24 P > F 0.031 0.163 0.016 Observation date [LS means ± 8.3 [LS means ± 2.7 (SE)] (SE)] First - 16.6b 2.9b Second - 20.5b 5.9b Third - 51.6a 27.4a F - 15.9 37.5 df - 2, 48 2, 54 P > F - <0.001 <0.001 *Means with the same letter in a column are not significantly different (Tukey adjustment, α = 0.05).

Figure 3-2. Sugarcane aphid infestation levels (LS means) in a sorghum field experiment evaluating seven biological insecticides at three post-treatment observation dates, spring 2019, Belle Glade, FL. Arrows represent insecticide applications.

CHAPTER 4 SUMMARY

The sugarcane aphid (Melanaphis sacchari) emerged as a sorghum (Sorghum bicolor) pest in North America in 2013 and became a concern in sorghum production in

Haiti in 2015. The development of resistant sorghum varieties has been the main approach for sugarcane aphid management in Haiti whereas in the United States a limited number of insecticides are used in combination with the adoption of resistant varieties. Hence, additional management tactics were studied to potentially provide a more comprehensive strategy for sugarcane aphid control.

Field experiments were conducted in Jonc-Labeille, Haiti in 2018 and 2019 to determine the effects of intercropping sorghum with maize (Zea mays) or pigeon pea

(Cajanus cajan) compared to a sorghum monoculture on sugarcane aphid infestations.

In both years, infestation levels were substantially lower in sorghum-maize than in sorghum-pigeon pea or sorghum alone. Relatively small sorghum plant size due to competition with maize plants likely contributed to the decrease in sugarcane aphid infestations in sorghum-maize. In addition, taller maize plants combined with sorghum plants during sorghum development may have acted as a barrier to sugarcane aphid colonization on sorghum.

The experimental plots were sampled for potential sugarcane aphid natural enemies using a standard 38-cm diameter sweep net. Sweep net sampling was complemented by direct observations. Aphid mummies were collected and maintained on sorghum leaves in Petri dishes in the laboratory at the American University of the

Caribbean in Les Cayes to observe adult parasitoid emergence. Coccinellids, chrysopids, syrphids, and braconids (Lysiphlebus testaceipes) were observed.

Plants of all grass species present in the experimental plots and within a 50-m- wide border area surrounding the experimental area were inspected for 1 to 3 hours once a week for identification of potential sugarcane aphid alternate hosts. Very low sugarcane aphid densities (1-4 aphids/infested plant) were observed on itchgrass

(Rottboellia cochinchinensis) throughout the sorghum growing season in the 2018 and

2019 experiments. An experiment with five potted itchgrass plants infested with five adult sugarcane aphids each was conducted twice to further investigate the potential of the plant as a sugarcane aphid host. The sugarcane aphids did not survive on the potted plants.

Azadirachtin, pyrethrins, Beauveria bassiana strain GHA, Isaria fumosorosea

Apopka strain 97, Chromobacterium subtsugae strain PRAA4-1T, Burkholderia spp. strain A396, and vetiver oil were compared to a conventional insecticide, flupyradifurone in the laboratory, greenhouse, and field. Flupyradifrone, azadirachtin, pyrethrins, B. bassiana, and the pre-mixed B. bassiana and pyrethrins negatively affected the aphid in the laboratory and greenhouse. However, C. subtsugae and vetiver oil negatively impacted the sugarcane aphid only in the laboratory. In contrast, sugarcane aphid infestations in insecticide-treated plots were not different than in non-sprayed plots in the field although flupyradifurone was consistently associated with the lowest infestations.

The results of this project suggest that intercropping sorghum with maize reduces sugarcane aphid infestations on sorghum. However, competition among maize and sorghum plants is a concern because of negative effects on sorghum yield. The results of this study also provide evidence that natural enemies of sugarcane aphid are present

in Haiti, which may provide some levels of control. Moreover, this study showed no evidence of sugarcane aphid colonizing grasses other than Sorghum spp. in Haiti. The results of this thesis project also suggest that biological insecticides including the pre- mix of B. bassiana and pyrethrins, pyrethrins, azadirachtin, and B. bassiana could potentially control sugarcane aphid infestations on sorghum if applied under favorable environmental conditions with good coverage. The use of these insecticides in a sugarcane aphid management program would allow reduced-risk sorghum production and insecticide resistance mitigation. The use of these insecticides would also be important in organic sorghum production.

LIST OF REFERENCES

Abate, T., A. van Huis, and J. K. O. Ampofo. 2000. Pest management strategies in traditional agriculture: an African perspective. Annu. Rev. Entomol. 45: 631-659.

Agboka, K., S. Gounou, and M. Tamò. 2006. The role of maize-legumes-cassava intercropping in the management of maize ear borers with special reference to Mussidia nigrivenella Ragonot (Lepidoptera: Pyralidae). Ann. Soc. Entomol. Fr. (n.s.). 42: 495-502.

Akmal, M., S. Freed, M. N. Malik, and H. T. Gul. 2013. Efficacy of Beauveria bassiana (Deuteromycotina: Hypomycetes) against aphid species under laboratory conditions. Pakistan J. Zool. 45: 71-78.

Altieri, M. A. 1999. The ecological role of biodiversity in agroecosystems. Agric. Ecosyst. Environ. 74: 19-31.

Amanullah, 2015. Competition among warm season C4 -cereals influence water use efficiency and competition ratios. Cogent Food Agric. 1:1011466. http://dx.doi.org/10.1080/23311932.2015.1011466

Amarasekare, K. G., and J. V. Edelson. 2004. Effect of temperature on temperature on efficacy of insecticides to differential grasshopper (Orthoptera: Acrididae). J. Econ. Entomol. 97:1595-1602.

Andon, J., and D. J. Sheltar. 2015. Curative control of hairy chinch bugs in lawn turf, 2013. Arthropod Manag. Tests: G1 (tsv177).

Araújo, J. P. M., and D. P. Hughes. 2016. Diversity of entomopathogenic fungi: which groups conquered the insect body? Adv. Genet. 94: 1-39.

Armstrong, J. S., W. L. Rooney, G. C. Peterson, R. T. Villenueva, M. J. Brewer, and D. Sekula-Ortiz. 2015. Sugarcane aphid (Hemiptera: Aphididae): host range and sorghum resistance including cross-resistance from greenbug sources. J. Econ. Entomol. 108: 576-582.

Armstrong, J. S., L. Mbulwe, D. Sekula-Ortiz, R. T. Villanueva, and W. L. Rooney. 2017. Resistance to Melanaphis sacchari (Hemiptera: Aphididae) in forage and grain sorghums. J. Econ. Entomol. 110: 259-265. ayiboPost. 2017. Des scientifiques Haïtiens en phase de sauver le petit mil. http://ayibopost.com/des-scientifiques-haitiens-en-phase-de-sauver-le-petit-mil/

Belhassen, E., J-J. Filippi, H. Brévard, D. Joulain, and N. Baldovini. 2015. Volatile constituents of vetiver: a review. Flavour Fragr. J. 30: 26-82.

Bianchi, F. J. J. A., C. J. H. Booij, and T. Tscharntke. 2006. Sustainable pest regulation in agricultural landscapes: a review on landscape composition, biodiversity and natural pest control. Proc. R. Soc. B. 273: 1715-1727.

Blackman, R. L., and V. F. Eastop. 1984. Aphids on the world’s crops: an identification guide. Wiley, Hoboken, NJ.

Bowling, R. D., M. J. Brewer, D. L. Kerns, J. Gordy, N. Seiter, N. E. Elliott, G. D. Buntin, M. O. Way, T. A. Royer, S. Biles, and E. Maxson. 2016. Sugarcane aphid (Hemiptera: Aphididae): a new pest on sorghum in North America. J. Integr. Pest Manag. 7: 1-13.

Brewer, M. J., R. Bowling, J. P. Michaud, and A. L. Jacobson. 2016. Sugarcane aphid: a new sorghum pest in North America. Texas A&M AgriLife Extension publication Ento-056, College Station, TX.

Brewer, M. J., J. W. Gordy, D. L. Kerns, J. B. Woolley, W. L. Rooney, and R. D. Bowling. 2017. Sugarcane aphid population growth, plant injury, and natural enemies on selected grain sorghum hybrids in Texas and Louisiana. J. Econ. Entomol. 110: 2109-2118.

Brown, S., D. Kerns, and J. Beuzelin. 2015. Sugarcane aphids: an emerging pest of grain sorghum. LSU AgCenter publication 3369, Baton Rouge, LA.

Buss, E. A., and S. G. Park-Brown. 2009. Natural products for insect pest management. University of Florida IFAS Extension publication ENY-350, Gainesville, FL.

Candy, J. 2008. Effet de la durée de compétition des mauvaises herbes sur la culture du poivron (Capsicum annuum). BS Thesis. Université Notre Dame d’Haiti, Port- au-Prince, Haiti.

Casida, J. E. 1980. Pyrethrum flowers and pyrethroid insecticides. Environ. Health Perspect. 34: 189-202.

Chandler, D., A. S. Bailey, G. M. Tatchell, G. Davidson, J. Greaves, and W. P. Grant. 2011. The development, regulation and use of biopesticides for integrated pest management. Phil. Trans. R. Soc. B. 366: 1987-1998.

Colares, F., J. P. Michaud, C. L. Bain, and J. B. Torres. 2015a. Recruitment of aphidophagous arthropods to sorghum plants infested with Melanaphis sacchari and Schizaphis graminum (Hemiptera: Aphididae). Biol. Control. 90: 16-24.

Colares, F., J. P. Michaud, C. L. Bain, and J. B. Torres. 2015b. Indigenous aphid predators show high levels of preadaptation to a novel prey, Melanaphis sacchari (Hemiptera: Aphididae). J. Econ. Entomol. 108: 2546-2555.

Cook, S. M., Z. R. Khan, and J. A. Pickett. 2007. The use of push-pull strategies in Integrated Pest Management. Annu. Rev. Entomol. 52: 375-400.

Copping, L. G., and J. J. Menn. 2000. Biopesticides: a review of their action, applications and efficacy. Pest Manag. Sci. 56: 651-676.

Cordova-Kreylos, A. L., L. E. Fernandez, M. Koivunen, A. Yang, L. Flor-Weiler, and P. G. Marrone. 2013. Isolation and characterization of Burkholderia rinojensis sp. nov., a non-Burkholderia cepacian complex soil bacterium with insecticidal and miticidal activities. Appl. Environ. Microbiol. 79: 7669-7678.

Davies, T. G. E., L. M. Field, P. N. R. Usherwood, and M. S. Williamson. 2007. DDT, pyrethrins, pyrethroids and insect sodium channels. IUBMB Life. 59: 151-162. de Souza, M. F., and J. A. Davis. 2019. Determining potential hosts of Melanaphis sacchari (Hemiptera: Aphididae) in the Louisiana agroecoscape. Environ. Entomol. 20: (in press).

Delgado-Ramírez, C. S., M. D. Salas-Araiza, O. A. Martínez-Jaime, R. Guzman- Mendoza, and S. Flores-Mejia. 2019. Predation capability of Hippodamia convergens (Coleoptera: Coccinellidae) and Chrysoperla carnea (Neuroptera: Chrysopidae) feeding on Melanaphis sacchari (Hemiptera: Aphididae). Fla. Entomol. 102: 24-28.

Diaz-Najera, J. F., S. Ayvar-Serna, J. C. Rodriguez-Maciel, M. Vargas-Hernandez, A. Mena-Bahena, and M. A. Tejeda-Reyes. 2018. Field efficacy of biorational insecticides against Melanaphis sacchari (Zehntner) in sorghum. Arch. Phytopathology Plant Protect. 51: 662-672.

Dua, V. K., A. C. Pandey, K. Raghavendra, A. Gupta, T. Sharma, and A. P. Dash. 2009. Larvicidal activity of neem oil (Azadirachta indica) formulation against mosquitoes. Malar. J. 8: 124.

Dubbs, A. L. 1971. Competition between grass and legume species on dryland. Agron. J. 63: 359-362.

Eckert, E., and A. Latané. 2017. The sorghum value chain in Haiti: a mapping and analysis. International Development Working Paper 2017-04, RTI International, Research Triangle Park, NC.

Edelson, J. V., J. Duthie, and W. Roberts. 2002. Toxicity of biorational insecticides: activity against the green peach aphid, Myzus persicae (Sulzer). Pest Manag. Sci. 58: 255-260.

Edwards, C. A, and R. S. Adams. 1970. Persistent pesticides in the environment. Crit. Rev. Env. Sci. Tec.1: 1-4.

Elliott, N., M. Brewer, N. Seiter, T. Royer, R. Bowling, G. Backoulou, J. Gordy, K. Giles, J. Lindenmayer, B. McCornack. and D. Kerns. 2017. Sugarcane aphid spatial distribution in grain sorghum fields. Southwest. Entomol. 42: 27-36.

Etheridge, B., J. Gore, A. L. Catchot, D. R. Cook, F. R. Musser, and E. J. Larson, 2019. Influence of temperature on the efficacy of foliar insecticide sprays against sugarcane aphid (Hemiptera: Aphididae) populations in grain sorghum. J. Econ. Entomol.112 :196-200.

Evans, G. A., and S. E. Halbert. 2007. A checklist of the aphids of Honduras (Hemiptera: Aphididae). Fla. Entomol. 90:518-523.

FAO. 2017. GIEWS - Global Information and Early Warning System. http://www.fao.org/giews/countrybrief/country.jsp?code=HTI

Fartek, B., S. Nibouche, P. Turpin, L. Costet, and B. Reynaud. 2012. Resistance to Melanaphis sacchari in the sugarcane cultivar R 365. Entomol. Exp. Appl. 144: 270-278.

Flecher, J. 2017. Distribution de semences de sorgho aux agriculteurs de la plaine du Cul-de-Sac. https://www.lenouvelliste.com/article/170717/distribution-de- semences-de-sorgho-aux-agriculteurs-de-la-plaine-du-cul-de-sac

Gabriel, S. 2016. Perte de 60% de la production de petit mil: une catastrophe annoncée pour Haiti! http://ayibopost.com/perte-de-60-de-la-production-de-petit-mil-une- catastrophe-annoncee-pour-haiti/

Gandarilla-Pacheco, F. L., M. S. Flores-Gonzáles, L. H. Morales-ramos, M. Elías- Santos, L. J. Galán-Wong, and I. Quintero-Zapata. 2015. Effect of native Mexican isolates of Isaria fumosorosea (Wize) Brown & Smith on Spodoptera exigua (Hübner) and Helicoverpa zea (Boddie). Southwest. Entomol. 40: 721- 729.

Ganyo, K. K., A. K. Tounou, C. Agboton, E. A. Dannon, B. R. Pittendrigh, and M. Tamò. 2012. Interaction between the aphid parasitoid Lysiplebus testaceipes (Hymenoptera: Aphididae) and its hyperparasitoid Syrphophagus africanus (Hymenoptera: Encyrtidae). Int. J. Trop. Insect Sci. 32: 45-55.

Hall, D. G. 1987. The sugarcane aphid, Melanaphis sacchari (Zehntner), in Florida. J. Am. Soc. Sugar Cane Technol. 7: 26-29.

Helden, M., and W. Tjallingii. 1993. Tissue localisation of lettuce resistance to the aphid Nasonovia ribisnigri using electrical penetration graphs. Entomol. Exp. Appl. 68: 269-278.

Henderson, G., R. A. Laine, D. O. Heumann, F. Chen, and B. C. R. Zhu. 2005a. Vetiver oil extracts as termite repellent and toxicant. US Patent No. 6,890,960 B1.

Henderson, G., D. O. Heumann, R. A. Laine, L. Maistrello, B. C. R. Zhu, and F. Chen. 2005b. Extracts of vetiver oil as repellent and toxicant to ants, ticks, and cockroaches. US Patent No. 6,906,108 B2.

Hunter, W. B., P. B. Avery, D. Pick, and C. A. Powell. 2011. Broad spectrum potential of Isaria fumosorosea against insect pests of citrus. Fla. Entomol. 94: 1051-1054.

IRAC [Insecticide Resistance Action Committee]. 2019. Mode of action classification scheme. Version 9.3. https://www.irac-online.org/documents/moa- classification/?ext=pdf

Jagtap, V., S. Bhargava, P. Streb, and J. Feierabend. 1998. Comparative effect of water, heat and light stresses on photosynthetic reactions in Sorghum bicolor (L.) Moench. J. Exp. Bot. 49: 1715-1721.

Jain, S. C., S. Nowicki, T. Eisner, and J. Meinwald. 1982. Insect repellents from vetiver oil: I. zizanal and epizizanal. Tetrahedron Lett. 23: 4639-4642.

Jones, G. A., and J. L. Gillett. 2005. Intercropping with sunflowers to attract beneficial insects in organic agriculture. Fla. Entomol. 88: 91-96.

K-State. 2016. USAID project led by Kansas State University to help develop new sorghum varieties for Haiti. http://www.hpj.com/crops/usaid-project-led-by-k- state-to-help-develop-new/article_3e5d6182-ade0-11e6-b0e5- eb3aa48cd434.html

Khan, Z. R., J. A. Pickett, J. van den Berg, L. J. Wadhams, and C. M. Woodcock. 2000. Exploiting chemical ecology and species diversity: stem borer and Striga control for maize and sorghum in Africa. Pest. Manag. Sci. 56: 957-962.

Khan, Z. R., C. A. O. Midega, A. Hassanali, J. A. Pickett, L. J. Wadhams, and A. Wanjoya. 2006. Management of witchweed, Striga hermonthica, and stemborers in sorghum, Sorghum bicolor, through intercropping with greenleaf desmodium, Desmodium intortum. Int. J. Pest Manag. 52: 297-302.

Khater, H. F. 2012. Prospects of botanical biopesticides in insect pest management. Pharmacologia. 3: 641-656.

Klingler, J., G. Powell, G. A. Thompson, and R. Isaacs. 1998. Phloem specific aphid resistance in Cucumis melo line AR 5: effects on feeding behavior and performance of Aphis gossypii. Entomol. Exp. Appl. 86: 79-88.

Knight, P. 1944. Insect associated with the palay rubber vine in Haiti. J. Econ. Entomol. 37: 100-102.

Knutson, A., R. Bowling, P. Porter, E. Bynum, R. Villanueva, C. Allen, and S. Biles. 2015. The sugarcane aphid. A new pest of grain and forage sorghum. Texas A&M AgriLife Extension, Texas A&M University, College Station, TX.

Kraiss, H., and E. M. Cullen. 2008. Insect growth regulator effects of azadirachtin and neem oil on survivorship, development and fecundity of Aphis glycines (Hemiptera: Aphididae) and its predator, Harmonia axyridis (Coleoptera: Coccinellidae). Pest Manag. Sci. 64: 660-668.

Kuhar, T. P., and H. Doughty. 2016. Evaluation of foliar and soil insecticides for the control of foliar insects in Summer squash in Virginia, 2015. Arthropod Manag. Tests. 41: tsw023.

Kumar, A. A. 2016. Botany, taxonomy and breeding, pp. 27-45. In Rackshit, S., and Y.- H. Wang (eds.), The sorghum genome, Compendium of Plant Genomes. Springer, Gewerbester, Switzerland.

Lama, L., B. E. Wilson, J. A. Davis, and T. E. Reagan. 2019. Influence of sorghum cultivar, phenological stage, and fertilization on development and reproduction of Melanaphis sacchari (Hemiptera: Aphididae). Fla. Entomol. 102: 194-201.

Lazzari, S., S. Starkey, J. Reese, A. Ray‐Chandler, R. McCubrey, and C. M. Smith. 2009. Feeding behavior of Russian wheat aphid (Hemiptera: Aphididae) biotype 2 in response to wheat genotypes exhibiting antibiosis and tolerance resistance. J. Econ. Entomol. 102: 1291-1300.

Leclerc, E., G. Pressoir, and S. Braconnier. 2013. L’avenir prometteur du sorgho sucré en Haiti. FACTS Reports [Online]. http://factsreports.revues.org/2801

Lee, D-H., B. D. Short, A. L. Nielsen, and T. C. Leskey. 2014. Impact of organic insecticides on the survivorship and mobility of Halyomorpha halys (Stål) (Hemiptera: Pentatomidae) in the laboratory. Fla. Entomol. 97: 414-421.

Levesque, A. 2014. Le sorgho sucré: une plante aux multiples valorisations, sources d’innovation en Haïti. BS. Thesis. Montpellier SupAgro, Montpellier, France.

Lipsey, B. A. Catchot, F. Musser, E. Larson, J. Gore, and D. Cook. 2017. Influence of planting population on sugarcane aphid (Melanaphis sacchari), pp. 680-684. In Proceedings, Beltwide Cotton Conferences, 4-6 January 2017, Dallas, TX.

Lowery, D. T., and M. B. Isman. 1994. Insect growth regulating effects of neem extract and azadirachtin on aphids. Entomol. Exp. Appl. 72: 77-84.

Maketon, M., N. Chakanya, N. Prem-udomkit, and C. Maketon. 2013. Interaction between entomopathogenic fungi and some aphid species in Thailand. Gesunde Pflanz. (Healthy Plants) 65: 93-105.

Marrone, P. G. 2019. Pesticidal natural products: status and future potential. Pest Manag. Sci. ps.5433.

Martin, P. A., D. Gundersen-Rindal, M. Blackburn, and J. Buyer. 2007. Chromobacterium subtsugae sp. nov., a betaproteobacterium toxic to Colorado potato beetle and other insect pests. Int. J. Syst. Evol. Microbiol. 57: 993-999.

Morehead, J. A., and T. P. Kuhar. 2017. Efficacy of organically approved insecticides against brown marmorated stink bug, Halyomorpha halys and other stink bugs. J. Pest Sci. 90: 1277-1285.

Muleta, K. T., G. Pressoir, and G. P. Morris. 2019. Optimizing genomic selection for a sorghum breeding program in Haiti: A simulation study. G3-Genes Genom. Genet. 9: 391-401.

Mulla, M. S., and T. Su. 1999. Activity and biological effects of neem products against arthropods of medical and veterinary importance. J. Am. Mosquito Contr. 15: 133-152.

[NPIC] National Pesticide Information Center. 2012. Neem oil. Oregon State University. Online. http://npic.orst.edu/factsheets/neemgen.html

[NPIC] National Pesticide Information Center. 2014. Pyrethrins. Oregon State University. Online. http://npic.orst.edu/factsheets/pyrethrins.html

National Pesticide Telecomunication Network. 1998. Pyrethrins and pyrethroids. Oregon State University. Online. https://www.cmmcp.org/sites/cmmcp/files/uploads/nptn_fact_sheet_on_pyrethrin s_pyrethroids.pdf

Natwick, E. T., and M. I. Lopez. 2015. Insecticide efficacy against pea aphid in alfalfa, 2014b. Arthropod Manag. Tests. 40: F28 (tvs157).

Nibouche, S., B. Fartek, S. Mississipi, H. Delatte, M. Reynaud, and L. Costet. 2014. Low genetic diversity in Melanaphis sacchari aphid populations at the worldwide scale. PLoS ONE. 9: e106067.

Nibouche, S. S. Mississipi, B. Fartek, H. Delatte, B. Reynaud, and L. Costet. 2015. Host plant specialization in the sugarcane aphid Melanaphis sacchari. PLoS ONE. 10: e0143704.

Nibouche, S., L. Costet, J. R. Holt, A. Jacobson, A. Pekarcik, J. Sadeyen, J. S. Armstrong, G. C. Peterson, N. McLaren, and R. F. Medina. 2018. Invasion of sorghum in the Americas by a new sugarcane aphid (Melanaphis sacchari) superclone. PLoS ONE 13: e0196124.

Niklova, I. M. 2016. Effects of organic products with insecticidal action on key insect pests in alfalfa seed production. Emir. J. Food Agric. 28: 609-615.

Pangnakorn, U. 2009. Efficiency of vetiver grass extracts against cowpea weevil (Callosopruchus maculatus Fabr.). American-Eurasian J. Agric. Eviron. Sci. 6: 356-359.

Ramanujam, B., R. Rangeshwaran, G. Sivakmar, M. Mohan, and M. S. Yandigeri. 2014. Management of insect pests by microorganisms. Proc. Indian Natn. Sci. Acad. 80: 455-471.

Reddy, G. V. P., and F. B. Antwi. 2016. Toxicity of natural insecticides on the larvae of wheat head armyworm, Dargida diffusa (Lepidoptera: Noctuidae). Environ. Toxicol. Pharmacol. 42: 156-162.

Roberts, D. W. 1989. World picture of biological control of insects by fungi. Mem. Inst. Oswaldo Cruz. 84: 89-100.

Rodriguez-del-Bosque, L. A., B. Rodriguez-Velez, M. A. Sarmiento-Cordero, and H. C. Arredondo-Bernal. 2018. Natural enemies of Melanaphis sacchari on grain sorghum in Northeastern Mexico. Southwest. Entomol. 43: 277-279.

Rodriguez-Saona, C. 2012. Can we make crops more attractive to the natural enemies of herbivores? Entomol. Ornitol. Herpetol. 1:1.

Rodríguez-Vélez, J. M., B. Rodríguez-Vélez, M. A. Sarmiento-Cordero, M. Palomares-Pérez, and H. C. Arredondo-Bernal. 2016. Species of Coccinellidae (Coleoptera: Cucujoidea) associated with Melanaphis sacchari Zehtner (Hemiptera: Aphididae) in Tamaulipas, Mexico. Entomol. News. 126: 97- 105.

Rodriguez-Vélez, J. M., A. Gallou, C. A. Uribe-Mu, M. A. Najar-Pacheco, F. M. Huerta-Martinez, A. Contreras-Ramos, and H. C. Arredondo-Bernal. 2019. Identification of twelve species of Coccinellidae (Coleoptera) predatory on Melanaphis sacchari (Zehntner) (Hemiptera: Aphididae) in Mexico, and submission of reference COI sequences. Coleopts. Bull. 73: 243-251.

Rubio, G., T. Walk, Z. Ge, X. Yan, H. Liao, and J. P. Lynch. 2001. Root gravitropism and below-ground competition among neighboring plants: a modeling approach. Ann. Bot. 88: 929-940.

Rundlöf, M., and H. G. Smith. 2006. The effect of organic farming on butterfly diversity depends on landscape context. J. Appl. Ecol. 43: 1121-1127.

Ruppel, R. F. 1983. Cumulative insect-days as an index of crop protection. J. Econ. Entomol. 76: 375-377.

Sabbour, M. M., and M. A. Abdel-Raheem. 2014. Evaluation of Isaria fumosorosae isolates against the red palm weevil Rhynchophorus ferrugineus under laboratory and field conditions. Curr. Sci. Int. 3: 179-185.

Safarzoda, S., C. A Bahlai, A. F. Fox, and D. A. Landis. 2014. The role of natural enemy foraging guilds in controlling cereal aphids in Michigan wheat. PLoS ONE. 9: e114230.

Salas-Araiza, M. D., D. R. López-Gutiérrez, O. A. Martínez-Jaime, and R. Guzmán- Mendoza. 2017. Parasitoids of sugarcane aphid, Melanaphis sacchari, at Irapuato, Guanajuato, México. Southwest. Entomol. 42:1091-1094.

Saranya, S., R. Ushakumari, S. Jacob, and B. M. Philip. 2010. Efficacity of different entomopathogenic fungi against cowpea aphid, Aphis craccivora (Koch). J. Biopest. 3: 138-142.

SAS Institute Inc. 2016. SAS/STAT 14.3 user’s guide. SAS Institute Inc., Cary, NC.

Selvaraj, K., and H. D. Kaushik. 2014. Greenhouse evaluation of Beauveria bassiana (Balsamo) Vuillemin against Aphis craccivora (Das) on Fenugreek. J. Appl. Nat. Sci. 6: 852-856.

Shapiro-Ilan, D. I., T. E. Cottrell, M. A. Jackson, and B. W. Wood. 2013. Control of key pecan insect pests using biorational pesticides. J. Econ. Entomol. 106: 257- 266.

Sharma, H. C., S. P. Sharma, and R. S. Munghate. 2013. Phenotyping for resistance to the sugarcane aphid Melanaphis sacchari (Hemiptera: Aphididae) in Sorghum bicolor (Poaceae). Int. J. Trop. Insect Sci. 33: 227-238.

Sharma, H. C., V. R. Bhagwat, D. G. Daware, D. B. Pawar, R. S. Munghate, S. P. Sharma, A. A. Kumar, B. V. S. Reddy, K. B. Prabhakar, S. S. Ambekar, and S. R. Gadakh. 2014. Identification of sorghum genotypes with resistance to the sugarcane aphid Melanaphis sacchari under natural and artificial infestation. Plant Breeding 133: 36-44.

Sheepmaker, J. W. A, and T. M. Butt. 2010. Natural and released inoculum levels of entomopathogenic fungal biocontrol agents in soil in relation to risk assessment and in accordance with EU regulations. Biocontrol Sci. Technol. 20: 503-552.

Singh, B. U., P. G. Padmaja, and N. Seetharama. 2004. Biology and management of sugarcane aphid, Melanaphis sacchari (Zehntner) (Homoptera: Aphididae), in sorghum: a review. Crop Prot. 23: 739-755.

Singh, D. 2014. Advances in plant biopesticides. Springer. Uttar Pradesh, India.

Smith, H. A., and R. McSorley. 2000. Intercropping and pest management: a review of major concepts. Am. Entomol. 46: 154-161.

Speight, M. R., M. D. Hunter, and A. D. Watt. 2008. Ecology of insects: concepts and applications. John Wiley & Sons Ltd. Chichester, UK.

Stark, J. D., and J. F. Walter. 1995. Neem oil and neem oil components affect efficacy of commercial neem insecticides. J. Agric. Food Chem. 43: 507-512.

Studebaker, G., and C. Jackson. 2017. Insecticidal control of sugarcane aphid, 2015. Arthropod Manag.Tests 42 (tsx079).

Szczepaniec, A., 2018. Interactive effects of crop variety, insecticide seed treatment, and planting date on population dynamics of sugarcane aphid (Melanaphis sacchari) and their predators in late-colonized sorghum. Crop prot. 109: 72-79.

Timyan, J. 1996. Bwa yo: important trees of Haiti. South-East Consortium for International Development, Washington, D.C., U.S.A.

Trenbath, B. R. 1993. Intercropping for the management of pests and diseases. Field Crops Res. 34: 381-405.

USAID/Haiti Mission-Wide. 2010. Pesticide Evaluation Report and safer Use Action Plan (PERSUAP).

Van den Berg, J., C. Midega, L. J. Wadhams, and Z. R. Khan. 2003. Can vetiver grass be used to manage insect pests on crops? pp. 254- 264. In Proceedings, third international vetiver conference, 6-9 October 2003, Guanzhou, China. The vetiver Network Award.

VanTine, M., S. Verlinden. 2003. Managing insects and disease damage under an organic system. Extension service, West Virginia University. https://organic.wvu.edu/files/d/f47962a9-d1e3-4bc8-a4d3- b7728548c30a/insects.pdf

Vedrine, E. W. 2003. Quelques plantes créoles et leurs noms scientifiques. http://www.potomitan.info/vedrine/kek_plant.php

Vesecky, J. F., K. C. Feltner, and R. L. Vanderlip. 1973. Wild cane and forage sorghum competition in grain sorghum. Weed Sci. 21:28-32.

Villanueva, R. T., M. Brewer, M. O. Way, S. Biles, D. Sekula, E. Bynum, J. Swart, C. Crumley, A. Knutson, P. Porter, R. Parker, G. Odvody, C. Allen, D. Ragsdale, W. Rooney, G. Peterson, D. Kerns, T. Royer, and S. Armstrong. 2014. Sugarcane aphid: a new pest of sorghum. Texas A&M AgriLife Extension publication Ento-035, College Station, TX.

Wall, G. C., and D. H. Meckenstock. 1992. Sorghum disease in Central America and the Caribbean basin, pp 67-73. In de Milliano, W. A. J., R. A. Frederiksen, and G. D. Bengston (eds), Sorghum and millets diseases : a second world review. ICRISAT, Patancheru, India.

Wang, N., L. Shi, D. Kong, D. Cai, Y. Cao, Y. Liu, G. Pang, and R. Yu. 2011. Accumulation levels and characteristics of some pesticides in human adipose tissue samples from Southeast China. Chemosphere 84: 964-971.

Waskom, R., T. Bauder, R. Pearson. 2017. Best management practice for agricultural pesticide use. Colorado State University Extension Bulletin XCM-177, Fort Collins, CO.

Webb, R. E., R. Peiffer, R. W. Fuester, K. W. Thorpe, L. Calabrese, and J. M. McLaughlin. 1998. An evaluation of the residual activity of traditional, safe, and biological insecticides against the gypsy moth. J. Arboric. 24: 286-293.

White, W. H., T. E. Reagand, and D. G. Hall. 2001. Melanaphis sacchari (Homoptera: Aphididae), a sugarcane pest new to Louisiana. Fla. Entomol. 84: 435- 436.

Wise, J. C., A. H. VanWoerkom, and R. Isaacs. 2015. Control of cranberry fruitworm in blueberry. Arthropod Manag.Tests. 40: C16 (tsv026).

Yadav, R. S. Prasad, S. K. Singh, V. Vijay, T. Sabu, S. Lama, P. Kumar, J. Sandesh, A. Thakur, and N. Ramawat. 2016. Bio-management of sugarcane aphid, Melanaphis sacchari (Z.) in sorghum. Plant Archives. 16: 559-562.

Zambrano-Gutierrez, J., R. Altorre-Rosas, M. G. Carillo-Benitez, J. R. Lomeli- Flores, R. A. Guzman-Plazola, A. Azuara-Dominguez, and A. P. Teran- Vargas. 2019. Species diversity of entomopathogenic fungi infecting the sugarcane aphid Melanaphis sacchari: a recently introduced pest in Mexico. Adv. Microbiol. 9: 38-55.

Zhu, B. C-R., G. Henderson, R. P. Adams, L. Mao, Y. Yu, and R. A Laine. 2003. Repellency of vetiver oils from different biogenetic and geographical origins against Formosan subterranean termites (Isoptera: Rhinotermitidae). Sociobiology 42: 623-637.

BIOGRAPHICAL SKETCH

Wilfrid Calvin was born in Lougou, a small community in southern Haiti. One year later his family and he moved to Les Cayes where he grew up and went to school until his undergraduate studies. In 2013, he completed a Bachelor of Science degree in agroforestry and environmental sciences at the American University of the Caribbean in

Les Cayes. During his studies, he had the opportunity to do two horticulture internships at Shroeder’s Flowers in Wisconsin, USA and Rockwell Farms in North Carolina, USA.

In 2014, he joined Frager Essential Oil SA as Fair-trade Internal Control Program

Coordinator. In 2016, he left this position to join the American University of the

Caribbean where he worked as Academic Assistant for 18 months.

In 2017, Wilfrid left behind his so loved wife and daughter to come to Florida to pursue his Master of Science degree at the University of Florida, Entomology and

Nematology Department, under the supervision of Dr. Julien Beuzelin. During his studies, Wilfrid addressed different management tactics to control the sugarcane aphid infesting sorghum in Haiti. Wilfrid conducted his research at the UF/IFAS Everglades

Research and Education Center and in Haiti. He completed his Master of Science degree in August 2019.