Toxicology

Insecticide Resistance Types and Mechanisms

1. W E L C O M E 1

2. INSECTICIDE RESISTANCE TYPES & MECHANISMS RESURGENCE Presented by: Ch. Naga Satyasri BAM-14-28 2

3. The added ability to withstand an insecticide, acquired by breeding of those individuals, which survive exposure to that particular insecticide insufficient to wipe out the whole colony. HOSKINS & GORDON (1956) Resistance to insecticide is the development of an ability in a strain of insects to tolerate doses of toxicant which would prove lethal to majority of individuals in a normal population of the same species. WORLD HEALTH ORGANISATION (W.H.O.) Definition 3

4. The first report of resistance came from Melander in 1914 who found sanjose scale, Quadrispidiotus sp . a pest of apples resistant to lime sulphur. Between 1916 and 1946, ten more cases of resistance to inorganic pesticides like arsenicals, selenium, cyanide and other compounds were reported. DDT which is patented in 1942, and by 1947, housefly resistance to DDT was documented. 4

5. After 1946, successive reports of insecticides resistance poured in the mosquito become resistant to DDT (1947) and housefly to pyrethrum (1952) and parathion (1952). By 1962, a list of 150 resistant insects becomes available and by 1976 the number went up to 300 insects. At present there are over 500 species of insects were found resistant to one or more insecticides. 5

6. Year Number of Resistant species 6

7. The occurrence of resistance to insecticide was first noticed in India in insects of public health importance. The first report of DDT resistance in mosquitoes came in 1952, in Culex fatigans a transmitter of filaria from U.P. and Mumbai. Since then this mosquito was resistant to both DDT and BHC in various parts of the country. 7

8. Resistance to agricultural pests was first reported by Pradhan et al ., (1963) on singhara beetle, Gelerucella birmanica in 1963 from Delhi against DDT and BHC. Srivastava and Joshi (1965) reported the BHC resistance in Spodoptera litura from Rajasthan. Beside BHC this pest was also developed resistance to Malathion , Endosulfan 8

9. The development of resistance in Tribolium castaneum to lindane and phospene ( Kalra et al .1975; Champ and Dyte , 1976). Orzaephilus surinamensis resistant to malathion and lindane (Champ and Dyte , 1976). In Khapra beetle, Dermestis granarium to phosphine (Borah and Chahal , 1979) has been reported in different parts of country. 9

10. Tribolium castaneum , Ephestia kuehniella and Plodia interpunctella shows resistance against spinosad ( Maedeh Mollaie , 2011 ). Tribolium castaneum shows resistance against the lufenuron ( Arora M.S ., 2012). 10

11. Planting of Bt crops globally and field-evolved resistance. 11

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14. Resistance is best understood as a process Initially, a small proportion of population has a resistant mechanism by chance. 14

15. The Resistance Process These individuals survive at a higher rate than others 15

16. Resistance as a process Resistant individuals increase in frequency 16

17. Resistance as a process Eventually, the pesticide or other management tactic causes too little control to be effective. 17

18. The process has three general stages, each with its own Management Strategy Prevention Manage or Reverse Abandon Pesticide/Management Tactic Need to monitor resistance 18

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20. Number of species of insects and mites reported resistant to insecticides Order Chemical group Cyclodienes DDT OP Carbmates Pyrethroids Fumigants Other Diptera 108 107 62 11 10 - 1 Lepidoptera 41 41 34 14 10 - 2 Coleoptera 57 24 26 9 4 8 5 Homoptera 15 14 30 13 5 3 1 Heteroptera 16 8 6 1 - - - Others 23 21 9 3 1 - 2 Acarina 16 18 45 13 2 - 27 Total 276 233 212 64 32 11 38 % 66 52 47 14 7 2 9 20

21. Examples of insects that tend to readily develop resistance to insecticides Insect Insecticides Resisted Green peach aphid Neonicotinoids , carbamates , organophosphates, pyrethroids , cyclodienes Diamondback moth Many insecticides Colorado potato beetle Many insecticides Maize earworm Pyrethroids , cyclodienes , carbamates , organophosphates Bed bugs Pyrethroids , cyclodienes , organophosphates Western root worm Cyclodienes , carbamates , organophosphates, Bt toxins Indian meal moth Organophosphates, cyclodienes , Bt toxins, pyrethrins Anopheles mosquitoes Many insecticides Tobacco budworm Many insecticides 21

22. How Resistance Develops……. Resistance usually develops by genetic mutations Types of mutations can include : A change in processes in the pest that make the pesticide harmless; A change in the place where the pesticide enters the pest so it cannot enter; A change in the behaviour of the pest so that it avoids the pesticide. 22

23. Insecticide resistance develops in target insects when a larva carrying a resistance mutation survives the application of a particular insecticide - breeds as a resistant insect. If a large proportion of the breeding population is then exposed to an insecticide of the same group selection for the resistance mutation will increase population of resistant insects will increase with time. 23

24. Source: McDougall, Sandra (2002) Integrated Pest Management in Lettuce: Information Guide. 24

25. Types of resistance Cross resistance Multiple resistance Negatively correlated resistance. 25

26. The rate of development of resistance in a population depends on….. The frequency of resistant genes present in a population The nature of genes (single or multiple; dominant or recessive) The intensity of the selection pressure of the toxicant The rate of breeding of the species 26

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28. Cross resistance It is operated when the same defence mechanism (usually governed by a single gene pair), such as increased oxidative detoxification, protects against several other insecticides. The phenomenon of cross resistance is a relatively frequent one in vector populations. For example, DDT and pyrethroid insecticides are chemically unrelated but both act on the same target site, the voltage gated sodium channel 28

29. Past use of DDT has resulted in several insect species developing resistance to DDT due to the kdr (Knock down resistant) mutation at the target site. Where these mutations have been retained in the population, the insects have some resistance to all pyrethroids in addition to DDT. Cross resistance can also occur between OP and carbamate insecticides when resistance results from altered AChE 29

30. Multiple resistance It occurs when resistance to various chemicals conferred by different coexisting defence mechanisms in the same insect strain, with each mechanism apparently coded by a different gene rather than all mechanisms controlled by a pleotropic expression of the same gene. 30

31. Localized populations of Colorado potato beetle are notorious for multiple resistance to more than 50 insecticides with various modes of action. 31

32. Negatively correlated resistance Selection of resistance by pressure from insecticide ‘A’ leads to a corresponding selection of susceptibility to insecticide ‘B’ vice versa. Thus, insect species which do not kill by ‘A’ can be controlled by ‘B’ for a few years, therefore alternate use of insecticide ‘A’ and ‘B’ can be utilized for pest suppression for a longer period. 32

33. Mechanisms of resistance There are several ways that insect populations can become resistant to insecticides, and pests may exhibit more than one of these mechanisms at the same time. Metabolic resistance is the most common type of resistance. Insects possess a variety of enzymes, including oxidases , glutathione S- transferases , esterases and amidases , whose function is to degrade foreign compounds. 33

34. Resistance to a toxicant can occur when the insects evolve an enhanced ability to detoxify or destroy the toxin, by alterations of these enzymes or of the amounts of enzymes produced in the body. As metabolic enzymes are directed at certain structural features of the toxicants, vulnerable sites in the molecule are affected. This type of resistance may not affect all members of a chemical class, and cross-resistance between compounds from different chemical classes can occur if they contain common chemical groups that are targeted by the same enzyme. 34

35. Metabolic resistance Resistant insects may detoxify or destroy the toxin faster than susceptible insects, or prevent the toxin from reaching target sites by binding it to proteins in their bodies. Metabolic resistance is the most common mechanism. Resistant insects may possess higher levels or more efficient forms of the enzyme(s) that break down insecticides to nontoxic compounds. 35

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37. Altered target-site resistance The site where the toxin usually binds in the insect becomes modified to reduce the insecticide's effects. Chitin is a major component of an insect's exoskeleton. 37

38. In the following example, enzyme H is necessary for chitin production. ( i ) To prevent moulting, the insecticide binds with the target site (ii) In a resistant insect, the target site is altered (iii) and prevents the insecticide from binding with the enzyme 38

39. Behavioral resistance   Resistant insects may avoid the toxin by a change from their normal activity.  Insects may simply stop feeding or move to the underside of a sprayed leaf. Some malaria-transmitting mosquitoes in Africa developed a preference for resting outside that prevented them from coming in contact with pesticide sprayed on interior walls.   39

40. Penetration resistance Resistant insects may absorb the toxin more slowly than susceptible insects. Penetration resistance occurs when the insect’s outer cuticle develops barriers which can slow absorption of the chemicals into their bodies.   This mechanism is frequently present along with other types.  40

41. Resistant insect Susceptible insect 41

42. Major biochemical mechanisms conferring resistance to important classes of insecticides in adult mosquitoes. Circle size reflects the relative impact of the mechanism on resistance 42

43. Major factors that influence resistance development Frequency of application : How often an insecticide or control tactic is used is one of the most important factors that influence resistance development. With each use, an advantage is given to the resistant insects within a population. 43

44. Dosage and persistence of effect : The length of time that an insecticide remains effective, also called its persistence, is dependent upon the physical chemistry of the insecticide, the type of formulation, and the application rate. Products which provide a persistent effect provide continual selection pressure in a similar manner to multiple treatments. It is important to always follow manufacturer and WHO recommendations when using insecticides. 44

45. Rate of reproduction : Insects that have a short life cycle and high rates of reproduction are likely to develop resistance more rapidly than species which have a lower rate of reproduction, as any resistance genes can rapidly spread throughout the population. Mosquitoes have a history of insecticide resistance and are characterised by a relatively short life cycle and high fecundity, with females laying several hundred eggs during their reproductive life. In contrast, the tsetse fly is less likely to develop resistance to insecticides due to a longer life cycle and relatively low rate of reproduction, females producing in total fewer than 10 offspring. 45

46. Insecticide resistance detection methods 1) Conventional Detection Methods 2)Biochemical detection of insecticide resistance 3) Immunological Detection Methods 4)Detection of monooxygenase ( cytocrome P450) based insecticide resistance. 46

47. Conventional Detection Methods Larvae or adults are tested for resistance by assessing their mortality after exposure to a range of doses of an insecticide. For susceptible and field populations, LD50 or LC50 values were calculated by using probit analysis. 47

48. Biochemical detection of insecticide resistance Biochemical assays/techniques may be used to establish the mechanism involved in resistance. When a population is well characterised some of the biochemical assays can be used to measure changes in resistance gene frequencies in field populations under different selection pressure.  48

49. Immunological Detection Methods:   This method is available only for specific elevated esterases in collaboration with laboratories that have access to the antiserum. There are no monoclonal antibodies, as yet, available for this purpose. An antiserum has been prepared against E4 carboxylesterase in the aphid Myzus persicae . An affinity purified 1gG fraction from this antiserum has been used in a simple immunoassay to discriminate between the three common resistant variants of M. persicae found in the UK field populations (Devonshire et al, 1996). 49

50. Detection of monooxygenase ( cytocrome P450) based insecticide resistance. The levels of oxidase activity in individual pests are relatively low and no reliable assay has been developed to measure p450 activity in single insects. The p450s are also a complex family of enzymes, and it appears that different cytocromes p450s produce resistance to different insecticides. 50

51. Impact of Resistance Overall agricultural productivity (during build phase) Increased pesticide usage Increased damage Environmental impact Increased pesticide usage Increased use of non-renewable resources Increased acreage Pest management flexibility Loss of pesticide tactic Constraint on new pesticides 51

52. Solution to the problem of resistance Occasional change of insecticide Use of undetoxifiable analogues Mixtures and alternations Negatively correlated insecticides Development of newer insecticides Use of insect pheromones Use of insect hormones Use of integrated approach Use of synergists 52

53. Judicious use of insecticide Indiscriminate use of insecticides lead to the development of resistance. Need based insecticidal spray should be taken up when pest population reaches the economic thresh-hold level. Selective insecticides should be sprayed using proper dose and right equipment. 53

54. Mixtures and Alteration of insecticide: One of the counter measure for resistance for one insecticide is to switch to another insecticide preferably belonging to different group. Examples There are reports stating DDT resistance in the housefly countered by substituting lindane or dieldrin I n India Heliothis armigera infesting cotton developed resistance against synthetic pyrethroids which could be countered by substituting acephate . 54

55. Use of undetoxifiable analouges : U se of two insecticides of independent action in a mixture against the resistant strain gives a good control and delays the development of resistance. Examples: I n cotton white flies developed against synthetic pyrethroids the mixture of cypermethrin and profenophos is used for overcoming the problem . DDT analogues such as prolan and Butalan (commercial mixture- Dilan ) lacking terminal chlorination on the central aliphatic chain are not easily detoxified by resistant strains. 55

56. Use of synergist Synergist enhances the toxicity of a given insecticide by inhibiting the detoxification mechanism . The first synergist was introduced in 1940 to increase the effectiveness of pyrethrum. The mode of action of synergist is the inhibition of mixed function oxidase enzymes that metabolize insecticides. Resistance to OP compounds, carbamates and pyrethrum are overcome by use of synergists such as piperonyl butoxide 56

57. Malathion resistance overcome by use of synergists such as triphenyl phosphate and tributyl phosphorotrithioate . Piperonyl butoxide and other methylene dioxyphenyl compounds serve to synergize many insecticides like DDT, carbamates etc., against houseflies and other insect pests. Propyl- paraoxon synergizes malathion and parathion through inhibition of phosphatase. 57

58. Negatively correlation insecticides Specific resistance to one insecticide leads to enhanced susceptibility to another insecticide. Cyclodiene resistant boll weevil have been found more susceptible to malathion 58

59. Development of newer insecticides: Switching over to newly evolved insecticide or to microbial insecticide is an important counter measure for delaying resistance. Use of insect pheromones Pheromone regulate insect behaviour and hormones regulate growth and reproduction. These can used successfully against resistance strains ; use of sex pheromones 59

60. Use of integrated approaches An integrated approach to control of insect pests will reduce the application of insecticides, which in turn, will lower insecticide pressure on insects. Under such condition genes governing resistance may not get activated or may take longer time to do so and keep resistance postponed for some time 60

61. RESISTANCE MANAGEMENT STRATEGY DESIGN Crop/Pest or Regional Strategies The strategies are provided on a CROP by PEST basis In many cases, a specific MoA insecticide can be used across this range of crops to control multiple pests that have the ability to move from crop to crop. The overall Resistance Management Strategy of avoiding overuse of individual Modes of Action insecticides should be followed, not just on a specific crop and pest but on a broad perspective of crops and pest complex. 61

62. Crop(s) : Sorghum, Maize, Summer Grain Legumes Insect(s) : Cotton bollworm/Native budworm ( Helicoverpa ) spp.) Monitor pest levels and do not spray unless pest thresholds are reached. Do not use consecutive applications of products from the same chemical group in Stages II and III . 62

63. In maize the critical stage of infestation- silking . Infestation could extend through to when the grain in cobs begins to harden although spraying for Heliothis is generally not required after silking is complete . In sorghum the critical stage of infestation-flowering and milky dough stage. Infestation could extend through to when the grain begins to harden . Post-harvest cultivation to destroy pupae, as practised in cotton crops, should also be practised in sorghum, maize and grain legume crops. 63

64. Conclusions : Resistance is caused by evolution — a fundamental biological process. The biochemical and physiological mechanisms may be many, but resistance is often due to an insensitive target for the pesticide or to increased detoxication . It is possible to curb or delay resistance development by making reproduction better for populations or subpopulations of pests that are not under selection pressure. 64

65. Cross-resistance may occur for pesticides with related modes of action, but sometimes also between nonrelated pesticides if they can be detoxified by the same enzyme. 65

66. Insecticide-resurgence 66

67. Definition Population after having been suppressed rebounds to numbers greater than before suppression occurred ( Pedigo , 1989 ) Increase in target arthropod ‘pest’ species abundance to a level which exceeds that of a control or untreated population following the application of an insecticide ( Hardin et al ., 1995 ) 67

68. It is defined as an abnormal increase of pest population often exceeding the economic injury threshold, following insecticide treatment ( Chelliah , 1979). Heinrich et al 1982 defined resurgence as a statistically significant increase in the insect population or damage in insecticide treated plots over that of untreated plots. 68

69. Historical perspective: The first instance of resurgence in rice was noticed in Brown plant hopper in the experimental fields of the IRRI. where the plots treated with gamma HCH @ 2 kg ai /ha recorded approximately three times higher population of BPH than the untreated control (IRRI, 1969). Resurgence has become a wide spread phenomenon in the past 2 decades ( Bhathal et al., 1991). 69

70. Mostly sucking insects are subjected to resurgence Their short life cycle Migrating ability Increased productivity and sensitiveness to the slight change in food and environment. 70

71. Crop Pests Resurgence causing pesticides Rice Brown planthopper (BPH) Perthane, endosulfan, acephate, phosphamidon, methomyl, cypermethrin, triazophos, decamethrin, diazinon, fenthion. Cotton Whiteflies DDT,triazophos, phosalone, synthetic pyrethroids like cypermethrin, decamethrin, fenvalerate Sugarcane Pyrilla, whitefly High level of nitrogenous fertilizers. Vegetables Chilli Yellowmite synthetic pyrethroids Lady’s finger Aphid Disulfotan, phorate, dimethoate Brinjal Fruit borer Acephate Muskmelon, watermelon Spidermite Ethion Pests, pesticides and resurgence in Indian context ( Jayaraj , 1987) 71

72. Causes of pest resurgence: Suppression of natural enemies: Insecticides, in general, exert adverse effects on the parasites and predators of crop pests and in many instances, this is considered as a major causes for insect pest resurgence to unmanageable proportions. Natural enemy mortality following broad spectrum insecticide application suggested as a important factor for BPH resurgence in rice (Miyashita, 1963) 72

73. Factors influencing pesticide – induced resurgence of insect pests 73

74. Pest Resurgence Pest (8) Natural Enemy 74

75. Pest Natural Enemy 75

76. pest pest 76

77. Pest Resurgence Note: 14 pests/leaf 77

78. Four processes contribute to resurgence Reduced Biological Control (Secondary) – most common with insects Reduced Competition – most common with weeds (mono vs. dicots ) Direct Stimulation of Pest – usually due to sub-acute doses Improved Crop Growth 78

79. Factors inducing resurgence Reduction in Natural Enemy Populations Direct effects Indirect effects Hormoligosis Alteration of plant quality Induction of insect detoxification enzymes by plant secondary chemicals and/or insecticides Direct and indirect enhancement of fecundity Effects of insecticide exposure on insect behaviour Reduction of competition 79

80. Preventing/Avoiding Pest Resurgence Avoiding hormoligosis Avoiding natural enemy destruction Avoiding competitor destruction Use physiologically selective control measures Use ecologically selective control measures Inoculative release of natural enemies Diversification of control methods 80

81. T h a n k Y o u !! 81