Pure Appl. Biol., 9(3): 1895-1902, September, 2020 http://dx.doi.org/10.19045/bspab.2020.90202

Research Article

Cross-negative effects of selective insecticides against different life stages of non-target pests, koenigii and hyalipennis on transgenic cotton under laboratory conditions

Waheed Ul Hassan1†, Talha Nazir1,2†*, Shah Zaman1, Tauqir Anwar1, Nawaz Haider Bashir1,3, Bushra Abid1, Muhammad Dildar Gogi1 and Muhammad Jalal Arif1 1. Department of Entomology, University of Agriculture Faisalabad, Faisalabad 38000-Pakistan 2. State Key Laboratory of Plant Diseases and Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193-China 3. Faculty of Plant Protection, Yunnan Agricultural University, Kunming, Yunnan 650201-China †These authors contributed equally to this work. *Corresponding author’s email: [email protected] Citation Waheed Ul Hassan, Talha Nazir, Shah Zaman, Tauqir Anwar, Nawaz Haider Bashir, Bushra Abid, Muhammad Dildar Gogi and Muhammad Jalal Arif. Cross-negative effects of selective insecticides against different life stages of non-target pests, Dysdercus koenigii and Oxycarenus hyalipennis on transgenic cotton under laboratory conditions. Pure and Applied Biology. Vol. 9, Issue 3, pp1895-1902. http://dx.doi.org/10.19045/bspab.2020.90202 Received: 01/02/2020 Revised: 25/04/2020 Accepted: 14/05/2020 Online First: 22/05/2020 Abstract Sucking insect pests, dusky cotton bug; Oxycarenus hyalinipennis Costa and red cotton bug; Dysdercus koenigii Walk has been great threat to transgenic (Bt) cotton worldwide. Present research was carried out to determine effect of some new chemistry insecticides [Steward® 150 EC (indoxacarb), Match® 50 EC (lufenuron), Coral® 36% SC (chlorfenapyr), Scatter® 15% SC (indoxacarb), Orchard® 13.3% EW (imidachloprid + abamectin) and Snap® 10% WDG (emamectin benzoate + tebufenozide)] under laboratory conditions (28 ± 2 °C, 65-70 % relative humidity and at photoperiod of 16:8 h of Light: Dark) to determine either field recommended dose rates have contact toxicity against non-target pests Dysdercus koenigii (: ) and Oxycarenus hyalipennis (: Hemiptera). Insecticides having contact mode of action that are recommended against chewing pest have also possess toxicity against sucking insect pests for which these pesticides are not recommended. Therefore the main objective of this research was to determine the contact toxicity against O. hyalinipennis and D. koengii. Laboratory bioassay results exhibit that Match® 50 EC was most toxic against nymphs, female and male of O. hyalinipennis and D. koengii and also showed maximum results in population reduction percentage for both under investigated pests. However, Orchard® has been caused minimum population reduction of both non-target pests among the tested insecticides. While remaining four insecticides Scatter®, Steward®, Coral® and Snap® showed moderate level of toxicity against tested insect pests. Keywords: Bioassay; Dysdercus koenigii; Insecticides; Life stages; Oxycarenus hyalinipennis; Toxicity

Published by Bolan Society for Pure and Applied Biology 1895 Hassan et al.

Introduction pests of cotton and crops, in South-east hirsutum L. (Cotton) is a main Asia and India as well [16]. In Egypt, oilseed and fiber crop [1]. Cotton has Dusky cotton bugs were reported as major significant contribution in earning through cotton pests, where it has caused weight foreign exchange. In Pakistan, cotton has losses in cotton seed and also caused been attacked by chewing and sucking decrease in oil seed quality and germination insect pests [2]. To control insect pest [17]. Dusky and Red cotton bugs feed on different methods are used especially seeds and unopened or opened bolls [18]. conventional chemical insecticides. When they crawls, a colored liquid excretes Globally Bt. Cotton is cultivated on large from bodies of these bugs on cotton lint scale to manage the bollworms population which results in staining like yellow spots [3, 4]. In Pakistan, chemical control is that ultimately reduces market value and considered as an immediate and effective quality of lint or textiles products. A control of insects pests [5-8]. Every year bacteria is also present in saliva that results about 80 % of the total agro-chemicals are in rottening of bolls and thus produced applied against insect pests of cotton that seeds from these damaged bolls has cause various problems [9-12]. germination and viability issues. Fungus With arrival of Bt cotton and changing in infestation occurs on the hole made due to sowing time season has effect the status of feeding of these bugs and results in higher all cotton pests. Sudden emergence of contents of the aflatoxin in the seed cake minor pest as a major or common pest is that is produced from such seeds which huge risk for cotton of Pakistan e.g.; cotton renders it not fit for the purpose of stink bugs and dusky cotton bugs [13, 14]. feeds [19]. First time dusky cotton bug (Oxycarenus Materials and methods hyalipennis) was reported in May, 2010 in For the determination of cross negative Israel, on Bt-Cotton and resulted in impacts of selective insecticides (Table 1), prematured fallings of the flowers along as contact toxic against two non-target with small bolls and squares / brackets. In insect pests, red cotton bug and dusky Pakistan as well as in several other cotton bug; research was performed in IPM countries of the world, Red cotton bug and (Integrated Pest Management) laboratory, Dusky cotton bug have gained the status of department of Entomology, University of pest. In Israel adults of the dusky cotton Agriculture Faisalabad, Faisalabad during bugs caused damages to fruit trees 2019. The research experiments were comprising grease spots on fruits as well as planned as these selected pesticides have cause deformity of the fruits [15]. In , not been examined before against these test dusky cotton bugs are known as major insects for their contact toxicity.

Table 1. List of the formulations of insecticides, chosen for current experiments Insecticides formulation Dose (gm or ml/acre) Match® 50 EC; (leufenuron) 250 Coral® 36 % SC; (chlorfenapyr) 150 Scatter® 15 % SC; (indoxacarb) 175 Orchard® 13.3 % EW; (abamectin+ imidachloprid) 200 Steward® 150 EC; (indoxacarb) 175 Snap® 10% WDG (tebufenozide+emamectin benzoate) 100

Collection of test insects; dusky cotton The population of red cotton bug and dusky bug and red cotton bug from infested cotton bug; adults and nymphs, was taken fields from different fields of cotton at University of Agriculture Faisalabad and taken to IPM

1896 Pure Appl. Biol., 9(3): 1895-1902, September, 2020 http://dx.doi.org/10.19045/bspab.2020.90202 laboratory of the University in order Steward® (indoxacarb) 150 EC, Snap® acclimatize the specimen with laboratory (tebufenozide+emamectin benzoate) 10% conditions. WDG and Coral® (chlorfenapyr) 36% SC Layout of Experiment were evaluated against selected test insect Different six insecticides; Scatter® pests. As per recommended doses for field (indoxacarb) 15% SC, Match® (lufenuron) application five concentrations of every 50 EC, Orchard® insecticide were formed (Table 2) by (abamectin+imidachloprid) 13.3% EW, applying Charles formula; C1V1=C2V2.

Table 2. List of concentrations applied for presented laboratory bioassays Concentrations Match® Coral® Scatter® Orchard® Steward® Snap®

T1 00.05 00.0625 00.04375 00.05 00.04375 00.025

T2 00.1 00.125 00.0875 00.1 00.0875 00.05

T3 00.2 00.25 00.0175 00.2 00.175 00.1

T4 00.4 00.5 00.35 00.4 00.35 00.2

T5 00.8 1 00.7 00.8 00.7 00.4 Control ------

Bioassay chewing insects pest showed cross negative For the purpose of bioassay of the selected impact on different non target insect pests. insecticides, method of filter paper All formulations of selected insecticides treatment was applied. Highest dose of cause inevitable population reduction of stock solutions were formed of the selected under investigated insect pests after 3 days insecticides with accurate dilutions. A of insecticides treatment (Table 3-8). scissor was used for the cutting of sterilized Among tested insecticides Match® 50 EC filter papers accordingly petri dish diameter (lufenuron) showed diverse impact and that was (90 x 15 mm). Then the dilutions cause considerable reduction in population of insecticides were applied by spraying on of Male, female and nymphs of under filter paper. Later on the treated filter paper investigated pests. Highest concentration were arranged in the petri dishes after (0.8%) caused reduction in population by drying. Filter paper treated with water was 86.18 (males), 81.76 (females), 85.61 arranged in petri dishes for control. For (numphs), 85.77 (males), 85.00 (females) each stage ten individuals (nymph, adult and 87.00% (numphs) for both insects (D. female and adult male) of both bugs were koenigii and O. hyalinipennis ) released at edges of the separated petri respectivelly. Mean mortalties of D. dishes with tender fresh cotton boll set at koenigii and O. Hyalinipennis gradually the center of every petri dish as bug food. increased with increase in concentration. All petri dishes were placed at 28 ± 2 °C During all these experiments, there has temperature and 65-70 % relative humidity been performed a comparative evaluation in laboratory. Three repeats were made for of some selective pesticides with different whole experiment with CRD. Data for mode of action and founded that Match® mortality was collected after three days of caused considerable and significant the post treatment interval. Henderson and reduction in population of non target insects Tilton formula was used for transforming of cotton (Table 3). These results shown mortality data into percent corrected that contact toxicity of Match® mortality [20]. (Leufenuron) 50 EC enhanced with the Results increase in the concentrations (Table 3). In vitro screening of different selective pesticides recommended against the

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Table 3. Means mortality of Dysdercus Koenigii and Oxycarenus hyalinipennis, Nymph, Male and Female exposed to various concentrations of Match® Mortality of D. koenigii Mortality of O. hyalinipennis Concentrations (Mean ± SE) (Mean ± SE)

Male Female Nymph Male Female Nymph 0.8 86.18A±0.55 81.76A±0.41 85.61A±0.29 85.77A±0.25 85.00A±0.91 87.00A±0.97 70.23B ± 58.69B± 69.00B ± 0.4 67.66B±0.69 62.53B±0.89 69.00B±0.88 0.69 0.89 0.87 43.30C ± 39.46C± 57.00C ± 0.2 52.5C ± 0.76 43.30C±0.81 45.00C±0.90 0.56 0.81 0.81 31.76D ± 24.07D± 37.00D ± 0.1 30.63D±0.83 27.92D±0.70 25.00D±0.73 0.83 0.70 0.69 17.00E ± 0.05 19.51E±0.69 8.69E ± 0.59 8.69E ± 0.51 7.33E ± 0.57 7.00E ± 0.56 0.44 Control 7.33F ± 0.39 8.33F ± 0.39 8.00F ± 0.32 4.84F ± 0.32 8.00F ± 0.38 5.00F ± 0.27 Different letters in column indicates statistical significance (at P ≤ 0.05) among the treatments

Second most toxic insecticide against non (nymphs) for both insects (D. koenigii and target insects was Coral®, where the O. hyalinipennis) respectivelly. Mean maximum concentration (1%) caused 72.42 mortality gradually decreased with (males), 67.66 (females), 86.85 (nymphs), decrease in concentration (Table 4). 66.51 (males), 72.42 (females) and 87.54%

Table 4. Means mortality of Dysdercus Koenigii and Oxycarenus hyalinipennis, Nymph, Male and Female exposed to various concentrations of Coral® Mortality of D. Koenigii Mortality of O. hyalinipennis Concentrations (Mean ± SE) (Mean ± SE) Male Female Nymph Male Female Nymph 72.42 A± 67.66 A± 86.85 A± 66.51 A± 72.42 A± 87.54 A± 1.0 0.81 0.79 0.75 0.87 0.93 0.86 51.00B ± 49.14B ± 58.14B ± 45.82B ± 58.14B ± 75.14B ± 0.5 0.66 0.57 0.87 0.61 0.81 0.76 40.28C ± 34.33C ± 40.28C ± 32.03C ± 43.85C ± 64.45C ± 0.25 0.76 0.83 0.76 0.76 0.75 0.51 29.57D ± 19.51D ± 26.00D ± 28.58D ± 40.28D ± 46.21D ± 0.125 0.57 0.91 0.63 0.51 0.73 0.48 15.28E ± 15.81E ± 15.28E ± 18.24E ± 26.00E ± 36.38E ± 0.625 0.61 0.64 0.55 0.45 0.67 0.41 4.66F ± 5.66F ± 5.33F ± 5.33F ± 5.66F ± 5.33F ± Control 0.49 0.51 0.39 0.34 0.47 0.37 Different letters in column indicates statistical significance (at P ≤ 0.05) among the treatments

In case of scatter®, the highest Scatter® (Indoxacarb) 15 % SC enhanced concentration (0.7%) cause reduction in with increase in concentrations (Table 5). population by 52.85 (males), 62.53 Statistical analysis showed that Orchard® (females), 57.00 (nymphs), 58.14 (males), was least toxic and caused minimum 54.84 (females) and 59.62% (nymphs) for population reduction of under investigation both insects (D. koenigii and O. insects among tested insecticides. hyalinipennis) respectivelly. Results have Maximum concentration of Orchard® also shown that the contact toxicity of (0.8%) caused 45.44 (males), 43.85 (females), 43.85 (numphs), 41.74 (males),

1898 Pure Appl. Biol., 9(3): 1895-1902, September, 2020 http://dx.doi.org/10.19045/bspab.2020.90202

40.28 (females) and 40.28% (nymphs) decrease use of such insecticides on reduction in population of examined insects transgenic cotton varieties increased the (D. koenigii and O. hyalinipennis) (Table occurrence of damage by minor pest and 5). Mean data showed that average ultimately those minor pests have gain the mortality of Orchard® was less than 50%. status of major pests of cotton crop. Earlier Moreover Steward® and Snap® caused 5% damage was reported by dusky cotton moderate reduction in population of tested bug on conventional varieties when mostly insects (Table 6-8). bolls of cotton got opened, consequently, Before extensive adaptation of genetically the injury produced by bugs was improved varieties of cotton, the insignificant. When peoples begun growing application of synthetic and conventional the Bt. Cotton initially; these bugs pyrethroids were considered the efficient developed very thoughtful danger for control against attack of bollworms on cotton crop [21]. conventional varieties of cotton. At present

Table 5. Means mortality of Dysdercus Koenigii and Oxycarenus hyalinipennis, Nymph, Male and Female exposed to various concentrations of Scatter® Mortality of D. Koenigii Mortality of O. Hyalinipennis Concentrations (Mean ± SE) (Mean ± SE) Male Female Nymph Male Female Nymph 52.85 A± 62.53 A± 57.00A± 58.14A± 54.84A ± 59.62A ± 0.7 0.55 0.79 0.94 0.89 0.96 0.78 38.03B ± 51.00B± 41.00B± 51.00B± 51.00B± 56.17B± 0.35 0.61 0.88 0.81 0.58 0.68 0.68 34.33C ± 31.76C ± 37.00C ± 40.28C ± 31.76C ± 45.82C ± 0.175 0.74 0.76 0.79 0.76 0.83 0.76 12.11D ± 24.07D ± 11.00D ± 22.42D ± 20.23D ± 32.03D ± 0.875 0.57 0.49 0.68 0.62 0.59 0.82 5.00E ± 8.69E ± 5.66E ± 8.69E ± 14.79E ± 0.4375 8.14E ± 0.56 0.49 0.53 0.51 0.41 0.65 1.00F ± 6.66F ± 0.66F ± 5.76F ± 5.33F ± Control 5.66F±0.41 0.38 0.59 0.47 0.37 0.51 Different letters in column indicates statistical significance (at P ≤ 0.05) among the treatments

Table 6. Means mortality of Dysdercus koenigii and Oxycarenus hyalinipennis, Nymph, Male and Female exposed to various concentrations of Orchard® Mortality of D. Koenigii Mortality of O. hyalinipennis Concentrations (Mean ± SE) (Mean ± SE) Male Female Nymph Male Female Nymph 45.44 A± 43.85 A± 43.85 A± 41.74 A ± 40.28 A± 0.8 40.28 A± 0.99 0.83 0.87 0.87 0.96 0.99 34.33B ± 36.71B ± 36.71B ± 34.33B ± 36.71B ± 0.4 36.71B ± 0.89 0.92 0.94 0.94 0.89 0.89 23.22C ± 26.00C ± 26.00C ± 23.22C ± 29.57C ± 0.2 29.57C ± 0.83 0.86 0.81 0.81 0.83 0.83 12.11D ± 18.85D ± 18.85D ± 19.51D ± 18.85D ± 0.1 18.85D ± 0.79 0.83 0.76 0.76 0.72 0.79 5.33E ± 6.00E ± 6.00E ± 12.11E ± 15.28E ± 0.05 15.28E ± 0.69 0.78 0.68 0.68 0.79 0.69 4.70F ± 4.57F ± Control 4.57F ± 0.56 6.33F ± 0.68 5.33F ± 0.52 5.33F ± 0.52 0.71 0.56 Different letters in column indicates statistical significance (at P ≤ 0.05) among the treatments

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Table 7. Means mortality of Dysdercus Koenigii and Oxycarenus hyalinipennis, Nymph, Male and Female exposed to various concentrations of Steward® Mortality of D. Koenigii Mortality of O. hyalinipennis Concentrations (Mean ± SE) (Mean ± SE) Male Female Nymph Male Female Nymph 49.14 A± 52.85 A± 72.51 A± 42.37 A ± 47.15 A± 0.7 33.00 A± 0.94 0.78 0.77 0.87 0.94 0.98 34.33B ± 41.74B ± 30.63B ± 32.03B ± 27.92B ± 0.35 17.00B ± 0.87 0.66 0.60 0.77 0.87 0.85 23.22C ± 26.92C ± 23.22C ± 28.58C ± 24.07C ± 0.175 9.00C ± 0.81 0.78 0.76 0.90 0.81 0.90 8.40D ± 19.51D ± 19.51D ± 18.24D ± 16.38D ± 0.875 5.00D ± 0.77 0.59 0.66 0.94 0.77 0.86 6.66E ± 5.33E ± 8.40E ± 5.66E ± 0.4375 7.89E ± 0.68 4.66E ± 0.68 0.55 0.78 0.75 0.79 1.00F ± 4.70F ± 6.33F ± 1.00F ± Control 5.00F ± 0.57 1.00F ± 0.57 0.48 0.57 0.67 0.63 Different letters in column indicates statistical significance (at P ≤ 0.05) among the treatments

Table 8. Means mortality of Dysdercus Koenigii and Oxycarenus hyalinipennis, Nymph, Male and Female exposed to various concentrations of Snap® Mortality of D. Koenigii Mortality of O. hyalinipennis Concentrations (Mean ± SE) (Mean ± SE) Male Female Nymph Male Female Nymph 33.14 A± 51.00 A± 52.72 A± 43.85 A± 47.15 A± 0.4 43.85A±0.543 0.80 0.83 0.74 0.83 0.96 26.00B ± 47.22B ± 38.93B ± 36.71B ± 31.76B ± 0.2 33.14B ± 0.47 0.63 0.71 0.85 0.76 0.89 15.28C ± 33.14C ± 32.03C ± 26.00C ± 24.07C ± 0.1 18.85C ± 0.39 0.76 0.76 0.79 0.70 0.76 11.71D ± 22.42D ± 28.58D ± 22.42D ± 12.53D ± 0.05 11.71D ± 0.76 0.68 0.69 0.62 0.76 0.67 4.57E ± 15.28E ± 18.24E ± 11.74E ± 0.025 4.57E ± 0.42 7.84E ± 0.61 0.57 0.51 0.57 0.68 3.66F ± 3.66F ± 5.00F ± 4.66F ± Control 3.33F ± 0.31 4.33F ± 0.53 0.49 0.43 0.46 0.55 Different letters in column indicates statistical significance (at P ≤ 0.05) among the treatments

The results of mean mortality indicates that impact on various life stages of the D. Match® exhibit maximum population koenigii and caused mortalties upto 96%. reduction of nymph, female and males of O. Against adults, Igr’s can result in hyalinipennis and D. koengii followed by malformation and produced deformed the Coral® and Snap®. However, Orchard® adults and nymphs as well as inhibit the showed minimum reduction in population wing growth. [24-26] founded that of tested insect pests that indicate that Lufenuron showed good chitin synthesis Orchard® was least toxic against various inhibiton effects against D. koenigii. Direct stages of life of O. hyalinipennis and D. application of Diflubenzuron inhibited koengii followed by Steward®. [22, 23] endocuticular deposition when applied on have been reported that Igr’s (Polyoxin D Mandusa epidermal. Three sites have been or diflubenzuron) showed maximum results proposed for describing the mode of action and are more efficacious against D. koenigii of diflubenzuron and other chitin synthesis and also noted that the Igrs cause negative inhibitors namely: inhibition of chitin

1900 Pure Appl. Biol., 9(3): 1895-1902, September, 2020 http://dx.doi.org/10.19045/bspab.2020.90202 synthetase (or its biosynthesis), inhibition fall armyworm and beet armyworm of proteases (or its biosynthesis) and injury. Proc Beltw Cott Conf, San inhibition of UDP-N-acetylglucosamine Diego, California, USA. 2: 1170-1172 transport through the membrane and these 5. Mohyuddin AI, Jilani G, Khan AG, findings contradict with our results as they Hamza A, Ahmad I & Mahmood Z have used other products. (1997). Integrated pest management of Conclusion major cotton pest by conservation, The overall findings of present study redistribution and augmentation of explained that selective insecticides caused natural enemies. Pak J Zool 29(3): normal to severe population reduction for 293-298. non-target sucking pests, which could lead 6. Gore J, Leonard BR & Adamczyk JJ to broad specturm resistance against (2001). Bollworm (Lepidoptera: different groups of pesticides. Pesticides Noctuidae) survival on ‘Bollgard’ and which cause death of non-selected pests ‘Bollgard II’ cotton flower bud and can cause major change to pest status as flower components. J Econ Entomol from non-pest to minorpest and from minor 94(6):1445-1451. to major pests. 7. Sarfraz M, Arif MJ & Ahmad G Authors’ contributions (2005). Comparative resistance of Conceived and designed the experiments: transgenic and conventional cotton W Hassan, T Nazir, MD Gogi & MJ Arif, against bollworms complex. Int J Agric Performed the experiments: W Hassan, T Biol 7(2): 308-310. Nazir & B Abid, Analyzed the data: W 8. Gogi, MD, Sarfaraz RM, Dosdall LM, Hassan, T Nazir & NH Bashir, Contributed Arif MJ, Keddie AB & Ashfaq M materials/ analysis/ tools: W Hassan, T (2006). Effectiveness of two insect Nazir, T Anwar & S Zaman, Wrote the growth regulators against Bemisia paper: W Hassan, T Nazir & S Zaman. tabaci (Gennadius (Homoptera: References Aleyrodidae) and Helicoverpa 1. Aslam M, Razaq M, Rana S, & Faheem armigera (Hubner (Lepidoptera: M (2004). Efficacy of different Noctuidae) and their impact on insecticides against bollworms on population densities of , cotton. J Res Sci 15(1): 17-22. predators in cotton in Pakistan. Pest 2. Nazir T, Gogi MD, Majeed MZ, Manag Sci 62: 982-990. Hassan W, Hanan A & Arif MJ (2017). 9. Bakhetia DRC, Singh J, Sohi AS & Field evaluation of selective systemic Singh J (1996). Integrated pest formulations against sucking insect management in sustainable agriculture: pest complex and their natural enemies Punjab scenario. Pest Manag Ecol Zool on a transgenic Bt-cotton. Pak J Zool 4: 1-13. 49(5): 1789-1796. 10. Ahmad M, Arif MI & Ahmad Z 3. Wilson FD, Flint HM, Deaton WR, (2000). Resistance of cotton whitefly, Fischoff DA, Perlak FJ, Armstrong Bemisia tabaci to cypermethrin, TA, Fuchs RL, Berberich SA, Parks NJ alphacypermethrin and & Stapp BR (1992). Resistance of zetacypermethrin in Pakistan. In: Proc. cotton lines containing a Bacillus Beltw. Cott. Conf., National Cotton thuringiensis toxin to pink bollworm Council, Memphis, TN. pp. 1015- (Lepidoptera: Gelechiidae) and other 1017. insects. J Econ Entomol 85: 1516- 11. Ahmad M, Arif MI and Ahmad Z 1521. (2001). Reversion of susceptibility to 4. Adamczyk JJ, Mascarenhas VJ, methamidophos in Pakistani Church GE, Leonard BR & Graves JB populations of whitefly, Bemisia (1998). Cotton boll susceptibility to tabaci. In: Proc. Beltw. Cott. Conf.,

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