Novel Approaches in Insecticide Resistance Management (IRM)

It gives elaborative description on Insecticide resistance, how it develops, mechanisms of insecticide resistance in insects, traditional and modern approach in resistance management

1. INSECTICIDE RESISTANCE MANAGEMENT IN INSECTS- NOVEL APPROACHES Presenting By: Suresh R. Jambagi 1 st Ph.D. (Entomology) PAMB0021 UAS, GKVK Bangalore 1

2. CONTENT 2 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

3. Insecticide Resistance? A heritable change in the sensitivity of a pest population that is reflected in the repeated failure of a product to achieve the expected level of control when used according to the label recommendation for that pest species. (IRAC) 3 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

4. Why Does Resistance Develop? Resistance is quick to develop to compounds with high effective kill, long residual and are highly selective at a single biochemical target site. Continued and frequent use of a insecticides Use of application rates -below or above those recommended on the label. Poor coverage of the area being treated Frequent treatment of organisms with large populations and short generation times. 4 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

5. 6. Failure to incorporate non-pesticidal control practices when possible 7. Simultaneous treatment of larval and adult stages with single or related compounds. 8. The toxicant is converted into a non toxicant form in the body of insect by various enzymes. 10. Genetic mutation and inheritance 5 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

6. Why Study Insecticide Resistance? A knowledge of insecticide resistance is an important from the point of view of proper selection of insecticides. 6 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

7. Resistance development in insects: Sl. no Year Insect Scientific name Insecticide 1 1914 San Jose scale Quadraspidiotus perniciosus Lime sulphur 2 1916 Citrus red scale Aonidiella aurantii HCN 3 1947 House fly Musca domestica DDT 4 1952 Mosquito Culex fatigans DDT 5 1952 Human louse Pediculus humanus corporis HCH 6 1953 Bed bug Cimex lectularius DDT 7 1961 Rat flea Xenopsylla cheopsis DDT 8 1963 Cattle tick Boophilus microplus Lindane 9 1963 Singhara beetle Galerucella birmanica DDT 10 1965 Tobacco caterpillar Spodoptera litura HCH 11 1971 Red flour beetle Tribolium castaneum DDT Patel et al ., 2018 7 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

8. Resistance across pest orders (IRAC) 8 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

9. World scenario: (IRAC) 9 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

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11. Mechanisms of insecticide resistance in insects 11 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

12. 1. Altered target-site resistance: The site where the toxin usually binds in the insect becomes modified to reduce the insecticide's effects. AchE insensitivity : Resistance to OP and Carbomates Sodium Channel insensitivity: Resistance to DDT and Pyrethroids GABA receptor (Cl-channel) insensitivity: Resistance to cyclodiens , fipronil, and avermectins . nAchR insensitivity: Resistance to nicotine, neonicotinoids and spinosad 12 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

13. 2. Behavioural resistance : Resistant insects may detect or recognize a danger and avoid the toxin by changing their normal activity . This mechanism of resistance has been reported for several classes of insecticides, including organochlorines, organophosphates, carbamates and pyrethroids. Examples: Anopheles mosquito: SS lives and bites inside home, but RR remains outdoor and flies into house to bite because DDT was applied to interior walls • House flies: Avoid treated surface • Cockroaches: Avoid treated surface and baits • Diamondback moth: Avoid permethrin 13 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

14. 3. Penetration resistance: It is due to modified composition and structure of integument Resistant insects may absorb the toxin more slowly than susceptible insects Binding protein and lipid reservoir traps insecticide in the cuticle. Penetration resistance is often present along with other forms of resistance. Slight resistance 14 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

15. 4. Metabolic resistance: METABOLISM: (Greek = Change) When an insecticide is applied on any living organism (plant/ animal), it undergoes some chemical changes resulting in the formation of new products called as metabolites and the process is metabolism. The extent and nature of the metabolites vary with the chemical, the organism ( species, strain, age, sex etc.), time and environmental factors. These metabolites may be more toxic, equitoxic or less toxic than original compound. 15 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

16. The normal enzymatic metabolism of insect is modified to increase insecticide detoxification or prevent activation of insecticides. Resistant insects may detoxify or destroy the toxin faster than susceptible insects or quickly rid their bodies of the toxic molecules. Resistant strains may possess higher levels or more efficient forms of these enzymes. Insects use their internal enzyme systems to break down insecticides. 16 Patel et al ., 2018 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

17. Esterase Resistance to OP, Carbamates and Pyrethroids Glutathione-S-Transferases R esistance to DDT, OP and Pyrethroids Cytochrome P 450 Mono- oxygeneses Resistance to all classes of insecticides Enzymes involved in the metabolic resistance mechanism 17 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

18. INSECTICIDE RESISTANCE MANAGEMENT (IRM): Goal Delay evolution of resistance in pests rather than its control/ management An effective insecticide resistance management program could be based on general resistance management principles endorsed by IRAC (Insecticide Resistance Action Committee). 18 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

19. Historically , growers respond to resistance by – 1. Increasing dosage and frequency of application which only accelerate the development of resistance. 2. Switch to another insecticide – so begins the pesticide treadmill . In theory, resistance can be minimized by – 1. Increase survivorship of Susceptible (SS) individuals. 2. Decrease survivorship of Resistant (R) individuals. 19 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

20. Figure: Graphic representation of the pesticide treadmill The pesticide treadmill is a term indicating a situation in which it becomes necessary for a farmer to continue using pesticides regularly because they have become an indispensable part of an agricultural cycle. 20 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

21. Principles of IRM 1. Management by Moderation 2. Management by Saturation 3. Management by Multiple attack (Curtis et al ., 1993) 21 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

22. 1. Management by Moderation: It attempts to preserve susceptibility genes are a valuable resource by limiting the chemical selection pressure that is applied. Infrequent applications Non-persistent chemicals Preservation of refugia Insect-resistant varieties Improved timing of planting and harvesting Encouragement of biological controls. 22 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

23. The term "saturation" does not imply saturation of the environment with pesticides. It is intended to indicate saturation of the insect's defenses by means of on-target dosages that are high enough to overcome resistance. This approach has more merit during the early stages of selection when resistance genes are rare, existing mainly in the heterozygous state. 2. Management by Saturation: 23 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

24. Formulations that could deliver high dosages on target include a. Microencapsulation b. Attractants c. Baited targets These will cause insecticide uptake at rates that are lethal to heterozygotes. 24 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

25. The multiple attack strategy is based on the premise that control can be achieved through the action of several independently acting stresses , including insecticides, each exerting selection pressure that is below the level which could lead to resistance. 3. Management by Multiple attack: 25 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

26. Achieved through: Alterations, sequences or rotations of compounds from group with different mode of action. Applications are often arranged into MOA spray windows or blocks that are defined by stage of crop development and biology of the pest concerned. Insecticide mixtures. Development of newer molecules with unique mode of action. 26 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

27. Concept Approach Means Moderation - Susceptible genes are a valuable resource that must be conserved while achieving economic control Low selection pressure Low doses producing < 100% mortality of SS Higher pest population thresholds for treatment Less frequent application Localized applications Preservation of refugia Some generations untreated Chemicals of short environmental persistence 2. Saturation - Removal of selective advantage of resistant phenotypes by saturation of defense mechanisms Rendering R genes functionally recessive Higher doses on target can render R genes functionally recessive, thus RS= SS Suppressing detoxification enzymes Appropriate synergist can cancel out specific detoxification enzymes and remove selective advantage of RS and RR 3. Multiple attack - Multi-directional, multi-site selection reduces degree of pressure by a single factor Maintaining degree of selection by each component factor below levels leading to resistance Mixtures of chemicals Rotation of chemicals Chemicals with multisite action 27 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

28. Insecticide Resistance Management Strategies Strategies to manage resistance are aimed at reducing the selection pressure from pesticide to a minimum while still achieving control. 28 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

29. Rotation of the insecticides Use of synergists Mixtures and alternations Negatively correlated insecticides Development of newer insecticides Use of insect pheromones Use of insect hormones Use of Integrated Approach (IPM) Spot spraying I. General IRM strategies: 29 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

30. 1. Rotation of the insecticides: The same insecticide should not used for a long time particularly when resistance has been detected. Insects resistant to one insecticide may not be resistant to another. After a time lag the lost susceptibility to the first insecticide returns 30 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

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32. Strategy YEAR 1 st Generation 2 nd Generation Strategy-I 1 st Year Intrepid 2 nd Year Avaunt, SpinTor , Danitol or Bt Strategy-II 1 st Year Intrepid Avaunt, SpinTor , Danitol or Bt 2 nd Year Avaunt, SpinTor , Danitol or Bt Intrepid Resistance insecticides Insecticides in IRM Guthion Imidan Intrepid Spintor Avaunt Danitol Bacillus thuringiensis Example: Tufted Apple Bud Moth 32 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

33. These inhibitory compounds we called synergists allow the insecticide to function normally only by blocking their inhibitory enzymes and do not in reality enhance the potency of the insecticides. Ex: Piperonylcyclone when mixed with DDT prevents its detoxification to DDE in the resistant strain house flies and these keeps them susceptible to DDT. As control agents, synergists can potentially render resistant populations susceptible and or prevent the development of resistance. 2. Use of Synergists: Synergist Insecticide DMC DDT ( Dehydrochlorinase ) PBO Pyrethroids Methylene dioxyphenyl Carbamates, Pyrethrin, DDT Propyl paraoxon Malathion, Parathion Gupta, 2001 33 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

34. Synergists are among the most straightforward tools for overcoming metabolic resistance because they can directly inhibit the resistance mechanism itself. It specifically interact with resistance associated with phase I metabolic enzymes (MFO). Use of PBO along with synthetic pyrethroids can delay the development of resistance in DBM and mosquitoes (Kumar et al ., 2002). NOTE: The WARF of the USA has developed a substance WARF antiresistant that can block DDT Dehydrochlorinase (the enzyme that detoxifies the DDT in houseflies ) and thus prevent the conversion of DDT to DDE and retain the insects susceptibility to this insecticide. 34 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

35. (Curtis et al ., 1993) 35 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

36. If an insect is treated with a synergist a few hours prior to exposure to an insecticide, it allows time for the synergist to cross the insect cuticle and inhibit those enzymes involved in metabolic resistance. It creates sensitivity or hyper sensitivity in insects. It reduces the insecticide dose to be subsequently applied. It can be achieved by - Split application of synergist and insecticide - Use of formulation technologies which allow a differential time release of synergist and insecticide Panini et al. , 2016 36 Temporal synergism concept Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

37. These increase the level of target pest control and broadening the range of pests controlled Insecticide mixtures may offer benefits for IRM when appropriately incorporated into rotation strategies with additional mode(s) of action, but generally a single mixture should not be relied upon alone. 3. Use of Mixtures: 37 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

38. Considerations: Should be compatible Individual insecticides selected for use in mixtures should be highly effective and be applied at the rates at which they are individually registered for against the target species. Mixtures with components having the same IRAC mode of action Classification are not recommended for IRM. When using mixtures, consider any known cross- resistance issues ‐ between the individual components for the targeted pest/s Mixtures become less effective if resistance is already developing to one or both active ingredients, but they may still provide pest management benefit 38 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

39. Ex: A BHC – DDT mixture against malaria mosquitoes (BHC is for DDT resistant survivors and DDT is for BHC resistant survivors) Note: A mixture of two insecticides and independent action has been suggested as a counter measure for the resistance. However in such cases it takes the a much shorter time for the insects to turn the resistant to both the compounds than what it would take if they are used separately 39 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

40. Resistance to one insecticide leads to the enhanced susceptibility to another insecticide. EX: 1. Cyclodiene resistant boll weevils are found to be susceptible to Malathion 2. DDT resistant houseflies also susceptible to Malathion Such combinations are discovered and used 4. Negatively correlated insecticides: 40 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

41. Just as pheromones modulate insect behaviour, hormones regulate growth and reproduction in insects. Both these processes can be interfered with by providing exogenous hormones at wrong times that is when they are not needed by the insect system . 5. Use of insect hormones in IRM: 41 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

42. It should be a continuous process to have more and more substitutions or alternates. There should be a constant attempt to search for newer insecticides This will make a large number of insecticides available for substitution when particular insecticides fails to kill 6. Developing newer compounds: 42 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

43. By treating border rows only (for pests that migrate into your planting from outside) and localized " hot spots " where pest numbers are over the action threshold. You leave large areas unsprayed for susceptible pests to survive. These areas can be treated at a later date, if thresholds are reached. 7. Spot spraying: 43 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

44. Pheromones regulate the insect behaviour. By using of the sex pheromones insects could be driven to poison baits and they will die. By providing the aggregation pheromones they could be driven to the wrong host plants where they would starve and die. To evaluate the insecticide resistance in the field 8. Pheromones Role: 44 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

45. An integrated approach to the control of insect pests will reduce the application of the insecticides which in turn will lower insecticides pressure on the insects under such a condition genes governing resistance may not get activated or may take a longer time to do so and keep resistance postponed for some time 9. INTEGRATED APPROACH: 45 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

46. Growers are requested to plant a non-Bt refuge around each plot of Bt cotton - Refuge must be at least 5 rows wide - The non-Bt refuge must be at least 20% of the total cotton plot The company supplies non-Bt seeds in a package attached to the Bt cotton seeds. - For each 450g Bt cotton seeds, 120g non-Bt cotton seeds Frequent monitoring of insect susceptibility is conducted II. Bt Crops: Refugia strategy 46 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

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48. Spatially separated applications of different compounds against the same insect constitute a “mosaic” approach to resistance management. By using two insecticides in different dwellings within the same village. This creates the potential for insects within a single generation to come into contact with both insecticides, and would reduce the rate of resistance selection, provided that multiple resistance within the vector population was extremely rare. If such a fine scale mosaic is to be used, careful records of which insecticide was used in each house are essential. III. Vectors and household pests: 1. Fine scale mosaic strategy 48 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

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50. ‘Control of pest population achieved by releasing large no. of sterile male insects, which will compete with normal males and reduce the insect population in subsequent generations’. E. F. Knipling (1937): Screwworm fly Methods of sterilization 1. Chemosterilants a. Alkylating agents- TEPA, Chloro ethylamine b. Antimetabolites- Amithopterin , 5-Fluororacil 2. Irradiation: X-rays, Neutrons,   2. Sterile Insect Release (SIRM) method: 50 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

51. After a decade of fighting for regulatory approval and public acceptance, Oxitec (A Firm from UK) released genetically engineered mosquitoes into the open air in the United States for the first time. Tests a method for suppressing populations of wild Aedes aegypti mosquitoes. Vector: Zika, dengue, chikungunya and yellow fever. Aim: Mate with the wild female population, responsible for biting prey and transmitting disease. The GM males carry a gene that passes to their offspring and kills female progeny in early larval stages. Male offspring won’t die but instead will become carriers of the gene and pass it to future generations. As more females die, the Aedes aegypti population should dwindle. Methodology: They placed boxes containing Oxitec’s mosquito eggs at six locations in three areas of the Keys. The first males are expected to emerge within the first two weeks of May (12000 males/week) Within 16 weeks: 20 million male mosquitoes will release to environment Oxitec mosquitoes carry a fluorescent marker gene (Nature, 2021) 51 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

52. Novel approaches in IRM Genome editing II. Genome editing in plants for resistance against insect pests Knocking down susceptible genes Modification of plant volatile blends Changing foliage colour Editing in insects for susceptibility toward plants/ insecticides Modification of Cry protein binding receptors Knockdown of detoxification enzymes. Editing of genes to disrupt chemical communication and mating partner identification Knock out of developmental genes Tyagi et al ., 2020 52 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

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54. I. Genome editing in insects It can be successfully employed in a two-step strategy involving the modification of target insects, and their subsequent release into the natural environment’ Modification of Cry protein binding receptors Knockdown of cadherin receptors that are genetically linked to Cry1Ac toxin resistance: Evidence for successful genome editing in H. armigera. T echni que - CRISPR/Cas9 system Target gene- HaCad This strategy can also be adapted to modify target sites in midgut receptors responsible for resistance development against BtICPs or insecticidal proteins. Wang et al ., 2016 54 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

55. 2. Knockdown of detoxification enzymes A. Knock-Down of Gossypol-Inducing Cytochrome P450 Genes Reduced Deltamethrin Sensitivity in Spodoptera exigua . Gossypol-fed beet armyworm larvae showed higher 7-ethoxycoumarin-O-deethylase (ECOD) activities and exhibited enhanced tolerance to deltamethrin. Meanwhile, gossypol-induced P450s exhibited high divergence in the expression level of two P450 genes: CYP6AB14 and CYP9A98 in the midgut and fat bodies contributed to beet armyworm tolerance to deltamethrin. Knocking down of CYP6AB14 and CYP9A98, via double-stranded RNAs (dsRNA) in a controlled diet, rendered the larvae more sensitive to the insecticide. These data demonstrate that generalist insects can exploit secondary metabolites from host plants to enhance their defense systems against other toxic chemicals. Impairing this defense pathway by RNA interference (RNAi) holds a potential to eliminate the pest’s tolerance to insecticides Hafeez et al ., 2019 55 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

56. B. CYP6AE gene cluster knockout in  Helicoverpa armigera  reveals role in detoxification of phytochemicals and insecticides Technique: CRISPR/Cas9 CYP6AE cluster contributes to the tolerance of two phytochemicals (xanthotoxin and 2-tridecanone) and two insecticides ( fenvalerate and indoxacarb). Fig. Responses to phytochemical toxins and insecticides Subfamily of CytP450 Wang et al ., 2018 The CRISPR-Cas9- based reverse genetics approach in conjunction with in vitro metabolism can rapidly identify the contributions of insect P450s in xenobiotic detoxification and serve to identify candidate genes for insecticide resistance. 56 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

57. 3. Editing of genes to disrupt chemical communication and mating partner identification A. Drosophila melanogaster Fruit flies are attracted by a diversity of odors that signal the presence of food, potential mates, or attractive egg-laying sites. In the case of Drosophila, a mutation in Or83b gene disrupted the selection of the egg-laying site (host) and impaired olfactory detection capacity . Or83b Encodes a Broadly Expressed Odorant Receptor Essential for Drosophila Olfaction Dendritic localization of conventional odorant receptors is abolished in Or83b mutants. Or83b mutation disrupts behavioural and electrophysiological responses to many odorants. Or83b therefore encodes an atypical odorant receptor that plays an essential general role in olfaction. Larsson et al ., 2004 57 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

58. B. Spodoptera litura Orco gene is an essential OR partner for both host and mate detection in Lepidoptera. Knock out of the Orco (olfactory receptor coreceptor) gene in Spodoptera litura caused distraction in mating partner selection and loss of identity toward host plants leaving them anosmia. Orco knockout caused defects in plant odor and sex pheromone olfactory detection in homozygous individuals Technique: CRISPR/Cas9 genome editing The CRISPR/Cas9 system induced Orco mutations at very high efficiency (89.6%) 80% of mutated individuals transmitted mutation to their progeny with up to 43.3% germline transmission efficiency. Koutroumpa et al., 2016 58 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

59. C. Helicoverpa armigera In insects, female adults release pheromones and convey males, the status of their maturity prior to mating. Males access the pheromone signals and select mature females. Knock out of odorant receptor 16 ( OR16 ) in H. armigera made males unable to receive pheromone signals from mature females. Results in mating with immature females that subsequently led to dumping of sterile eggs. Technique: CRISPR/ Cas9 Knock out of OR16 receptor in lepidopteran pests can therefore be a novel and effective strategy to regulate mating time for pest management in agricultural crops. Sun et al ., 2017 59 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

60. 4. Knock out of developmental genes Knocking out developmental genes such as abd -A (Abdominal A) gene, a transcriptional factor involved in downstream regulation of various target genes that are extensively involved in development. Loss of function mutants through CRISPR/ Cas9 resulted in the generation of abd -A mutant phenotypes in various agricultural pests like Spodoptera litura , Spodoptera frugiperda and Plutella xylostella . Insects thus produced showed deformity in body segments, disarmed prolegs, anomalous gonads, and embryonic lethality indicating the success of gene editing tools . Target insect Target gene Reference Spodoptera litura Sl abd -A Sun et al ., 2017 Spodoptera frugiperda Sf abd -A Wu et al ., 2018 Plutella xylostella abdominal-A Sun et al ., 2017 60 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

61. Background: Editing plants for insect pest management has been less exploited so far (Tyagi et al ., 2020). II. Genome editing in Plants Base: Most polyphagous pests recognize host plants using the plant’s own volatile blends, visual appearance, gustatory clues, oviposition sites, and their interactions are coevolved. 61 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

62. 1. Knocking down susceptible genes Emerging as a reliable strategy Insects rely on important chemical components from plants for their development, immunity, and behavior. In rice, CRISPR/Cas9-based knock down of CYP71A1 gene encoding tryptamine 5- hydroxylase which catalyzes the conversion of tryptamine to serotonin resulted in reduced growth in brown plant hopper. The study was hypothesized on the fact that serotonin, a neurotransmitter from plants is essential for larval immunity and behavior. Lu et al ., 2018 62 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

63. 2. Modification of plant volatile blends Studies have demonstrated that changes in volatile blends retract insects from host plants. In transgenic Arabidopsis thaliana plants, aphid ( Myzus persicae ) infestation leads to release of a sesquiterpene hydrocarbon (E)-β-farnesene (Eβf) which retracts feeding by other host populations and attracts a parasitic wasp Diaeretiella rape that controls aphid population. Note: Care are should be taken such that the modification will not lead to deleterious effects toward beneficial insect population. Beale et al ., 2006 63 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

64. 3. Changing foliage colour Visual appearance of plants plays a prominent role in the ability of insects to recognize and attack host plants. Alteration in plant pigmentation has been found to modify insect host preferences. Red leaf tobacco: Transgenic- Overproduction of anthocyanin pigmentation- red coloration of leaves This change in leaf color proved to be acting as a deterrent to the herbivores, Spodoptera litura and Helicoverpa armigera confirming the importance of leaf color and appearance on host recognition in insects. Upon feeding S. litura - Delayed pupation H. armigera- Increase in larval mortality Malone et al ., 2009 64 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

65. Challenges to Managing Resistance Do not have adequate tools Not enough registered products to permit rotation The program may not be as effective The program may be more expensive Products with resistance risk for one pest are also used for others 65 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

66. CONCLUSION 66 Suresh R. Jambagi Ph.D. (Entomology) UAS GKVK Bangalore

67. THANK YOU ALL… 67