Pak. Entomol. Vol. 32, No.

2, 2010

PAKISTAN ENTOMOLOGIST ISSN 1017-1827 http://www.pakentomol.com email: pakentoeditor@gmail.com

EVALUATION OF INSECTICIDE RESISTANCE IN TWO STRAINS OF FRUIT FLY, BACTROCERA ZONATA (SAUNDERS) (TEPHRITIDAE: DIPTERA), WITH FRUIT DIP METHOD
Syed Faisal Ahmad, Sohail Ahmed, Rashad Rasool Khan and Muhammad Kashif Nadeem Department of Agri. Entomology, University of Agriculture, Faisalabad, Pakistan. ABSTRACT Present study was conducted to determine the level of insecticide resistance in five insecticides, viz., malathion, trichlorfon, lambda-cyhalothrin, spinosad and bifenthrin, commonly being used for the control of fruit flies, in two field strains of fruit fly, Bactrocera zonata (Saunders) (Tephritidae: Diptera) by using fruit dip bioassay method under laboratory conditions. Response data were collected at 24 and 48 hours after releasing graved females in the cages having fruits dipped in appropriate concentrates of each insecticide. Results showed that B. zonata from Multan and Faisalabad zones were resistant to trichlorfon, malathion, lambda-cyhalothrin and bifenthrin ranging 3-19 fold, however, population from these places were susceptibility to spinosad. Malathion registered resistances ratio (3-6 fold) less than bifenthrin (8-11 fold), trichlorofon (10-19 fold) and cyhalothrin (4-9 fold). The data suggest that B. zonata has developed resistance to trichlorfon and indicate a danger for its use as cover spray and in baits. Keywords: Fruit fly, Bactrocera zonata, insecticide, resistance. INTRODUCTION About 11 species of fruit flies are recorded from Pakistan and most notable among them are Bactocera zonata, B. cucurbitae, B. dorsalis, Myiopardalis pardalina, Carpomiya incompleta, C. vesuviana, Dacus ferrugincus and D. diversus (Abdullah and Latif, 2001; Abdullah et al., 2002; Stonehouse et al., 2002; Panhwar, 2005). The hosts of fruit flies recorded in Pakistan are apple, ber, guava, mango, musk melon and bitter gourd (Khan and Musakhel, 1999; Sultan et al., 2000; Ahmad et al., 2005). Fruit flies rest on non-host plants which may or may not sprayed. If not sprayed directly, these might have residues of insecticides by means of drifts from fields where insecticides have been applied. The baiting which also includes insecticides is another source of fruit flies succumbed to insecticide pressure. These two reasons are enough to justify speculation about insecticide resistance in fruit flies. The topical method of insecticide application was used to analyze the resistance in laboratory bioassay which has been a standardized method with Food and Agriculture Organization (FAO) as well. Other methods of application have received less attention and fruit dip is one of these methods (Hsu and Feng, 2000; Maklakov et al., 2001). The extent of damage reported by the fruit flies specie, Bactrocera dorsalis was 5-100% loss in Pakistan (Syed et al., 1970). Damage caused by fruit flies to fruit and vegetable growers in Pakistan is about 200 million US dollars annually at farm level with added losses to traders, retailers and exporters (Stonehouse et al., 1998). The greatest threat caused by the fruit flies is the rejection of fruit commodity especially mangoes due to presence of its maggots and make it unfit for human consumption (Stonehouse et al., 2002). In Pakistan, the control of fruit flies is mainly dependent on the use of insecticides. These insecticides have different methods of application such as baiting, attractant insecticides and cover sprays. The heavy infestation of fruit flies has lead to

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Pak. Entomol. Vol. 32, No.2, 2010 the use of cover sprays (Anonymous, 1986). The insecticides, for example, organophosphates, carbamates, synthetic pyrethroids and new chemistry are being indiscriminately used by farmers as cover sprays (Stonehouse et al., 1997; Alston, 2002; Ahmad et al., 2005; El-Aw et al., 2008). In this paper, we have determined resistance in some populations of B. zonata collected from Multan and Faisalabad and compared with a laboratory reared population against bifenthrin (Talstar 10 EC), trichlorfon (Diptrex 80 SP), lambda-cyhalothrine (Karate 2.5 EC), Malathion (Fyfanon 57 EC) and spinosad (Tracer 240 SC). MATERIALS AND METHODS Present study was conducted under laboratory conditions (28oC, 555% RH) in Department of Agri. Entomology, University of Agriculture, Faisalabad to determine the level of resistance against five insecticides against fruit fly, B. zonata (Saunders) commonly used for its control. Field populations of B. zonata from Faisalabad and Multan zone were collected from the infested and fallen fruits of the mangoes and guava. A reference susceptible strain was obtained from CABI, Multan in pupal form which has been reared for the last many years without insecticide exposure for comparison. The experiment was laid down according to completely randomized design with three replicates including an untreated control. Rearing of both susceptible and field strains of B. zonata were carried in Perspex cages on healthy mango fruits for egg laying. The infested fruits were then placed in a wooden cage having soil at the bottom for pupation. The pupae were isolated from the soil and placed in a separate cage for emergence. The adults were fed on banana based artificial diet having ingredients such as egg yolk, sugar, honey, yeast, syrup vitamin B complex blended in the ratio of 2:4:8:2:2:1 respectively in an electric blender to make a paste and was kept in a freezer for subsequent use. Commercial formulations of tested insecticides were obtained from their manufacturers and were used after their dilutions in distilled water for the determination of level of resistance against field populations of B. zonata. 5-6 serial concentrations of each insecticide were made to obtain most practical concentration that yielded mortality between 5-95% in each case. Fruit dip bioassay method was used to expose both susceptible and field strains of B. zonata against insecticides. Fresh mangoes were dipped in the insecticide solution for 2-3 seconds and then were air dried. Treated fruits were placed in the plastic jars and equal numbers of 12-15 days old female adults were transferred to each jar where they were also provided with the normal adult diet. The openings of the plastic jars were covered with muslin cloth in order to prevent escape of flies and also for proper aeration. Irreversible knockdown followed by death of the adult fruit flies was the criterion to determine mortality at the intervals of 24 and 48 hours. Data for adult mortality were subjected to Probit Analysis and was corrected by Abbotts formula (Abbott, 1925) and analyzed by Probit Analysis (Finney, 1971) using the software POLO-PC (LeOra Software, 1987). The LC50 values of each compound were estimated and compared with that of the laboratory strain to determine the ratio of insecticide resistance. RESULTS Response of B. zonata strains against bifenthrin LC50 (ppm) of bifenthrin against Faisalabad population was 458.62 at 24 hrs which was reduced to 247.77 after 48 hrs. In susceptible strain the value of LC50 was found to be 55.79 and 25.2 ppm after 24 and 48 hrs, respectively. Thus, Faisalabad strain exhibited the resistance ratio of 8 folds after 24 hrs with an increase up to 9.8 folds after 48 hrs. In Multan strain, LC50 of bifenthrin was found to be 533.33 and 279.28 ppm at 24 and 48 hrs, respectively, with the resistance ratio of 11 folds (Table 1). Response of B. zonata strains against trichlorfon Faisalabad strain was found to be 10 and 13 folds more resistant as compared to the susceptible strain with the LC50 (ppm) of 133.33 and 61.93 at 24 and 48 hr respectively in comparison to that of susceptible strain 12.79 and 4.75 at 24 and 48 hrs, respectively (Table 2). Multan strain was found more resistant (69) to trichlorofon with the LC50 values of 214.45 and 93.67 ppm at 24 and 48 hrs, respectively (Table 2). Response of B. zonata strains against malathion LC50 (ppm) for Faisalabad strain, respectively, were 60.81 and 24.96 ppm at 24 and 48 hrs (Table 3) with low resistance ratio of 3 folds.

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Pak. Entomol. Vol. 32, No.2, 2010 Table 1. LC50 and Fit of Probit lines in strains of B. zonata against bifenthrin. CI Fit of Probit line Observation LC50 Strains (hrs ) (ppm) (95%) SlopeSE 2 24 55.79 39.16-91.63 1.410.27 0.38 Laboratory 48 25.2 18.17-33.55 1.90.31 1.29 24 458.62 305.89-852.04 1.210.26 0.22 Faisalabad 48 247.77 172.93-345 1.720.31 1.12 24 533.33 367.8-950.23 1.380.27 0.90 Multan 48 279.28 168.78-331.91 1.760.31 0.96 CI= Confidence interval; RR= Resistance ratio; LC50= Lethal concentration.

RR 8.22 9.83 9.55 11.08

Table 2.-LC50 and Fit of Probit lines in strains of B. zonata against trichlorfon. CI Fit of Probit line Observation LC50 Strains (hrs ) (ppm) (95%) SlopeSE 2 24 12.79 8.86-21.11 1.340.27 0.52 Laboratory 48 4.75 3.34-6.27 1.980.33 2.92 24 133.33 91.95-237.56 1.380.27 0.90 Faisalabad 48 61.93 42.86-86.83 1.680.31 0.91 24 214.45 151.67-343.31 1.440.27 0.20 Multan 48 93.67 64.84-127.33 1.730.30 0.84 CI= Confidence interval; RR= Resistance ratio; LC50= Lethal concentration.

RR 10.42 13.03 16.76 19.72

Table 3. LC50 and Fit of Probit lines in various strains of B. zonata against malathion. Observation LC50 CI Fit of Probit line Strains (hrs ) (ppm) (95%) SlopeSE 2 24 16.66 11.49-29.69 1.380.27 0.90 Laboratory 48 7.74 5.4-7.78 1.720.31 1.12 24 60.81 41.94-106.4 1.350.27 0.17 Faisalabad 48 24.96 16.93-34.45 1.710.31 0.99 24 91.68 64.35-142.98 1.40.27 0.10 Multan 48 48.17 33.46-65.7 1.730.30 0.97 CI= Confidence interval; RR= Resistance ratio; LC50= Lethal concentration.

RR 3.65 3.22 5.50 6.22

Table 4. LC50 and Fit of Probit lines in strains of B. zonata against lambda-cyhalothrin. Observation LC50 CI Fit of Probit Line Strains (hrs ) (ppm) (95%) SlopeSE 2 24 53.61 37.91-85.82 1.440.27 0.20 Laboratory 48 17.49 7.72-28.38 2.000.33 3.36 24 229.97 161.47-380.43 1.420.27 0.21 Faisalabad 48 84.91 61.88-110.1 2.280.37 1.99 24 422.44 229.84-703.39 1.350.27 0.56 Multan 48 162.13 113.84-214.50 1.420.27 2.87 CI= Confidence interval; RR= Resistance ratio; LC50= Lethal concentration.

RR 4.28 4.85 7.87 9.26

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Pak. Entomol. Vol. 32, No.2, 2010 Table 5. LC50 and Fit of Probit lines in strains of B. zonata against spinosad. CI Fit of Probit Line Observation LC50 Strains (hrs ) (ppm) (95%) SlopeSE 2 24 53.61 37.91-85.82 1.440.27 0.20 Laboratory 48 19.89 13.92-26.6 1.870.31 0.87 24 41.73 28.85-65.02 1.350.26 0.23 Faisalabad 48 21.61 15.48-28.35 2.140.35 2.5 24 87.45 59.46-141.86 1.280.26 0.76 Multan 48 36.96 26.51-41.08 2.280.37 1.29 CI= Confidence interval; RR= Resistance ratio; LC50= Lethal concentration. LC50 for the Multan strain was 91.68 ppm at 24 hrs which was decreased to 48.17 ppm after 48 hrs of exposure with resistance ratio of 5 and 6 fold, respectively. Response of B. zonata strains against lambdacyhalothrin LC50 (ppm) for lambda-cyhalothrin was 53.61 and 17.49 at 24 and 48 hrs with RR of 4 folds. LC50 of 422.44 (7 folds) and 162.13 (9 folds) ppm was recorded in Multan strain (Table 4). Response of B. zonata strains against spinosad Both the field populations of B. zonata were found highly susceptible to spinosad (Table 5). After the exposure of 24 hrs, resistance ratio in Faisalabad strain was found to be 0.77, which increased to 1.08 after 48 hrs presenting susceptibility with the LC50 (ppm) of 41.73 and 21.61 after 24 and 48 hrs, respectively, whereas, Multan strain exhibited the resistance ratio of 1 both after 24 and 48 hrs, and also showed susceptibility towards spinosad with the LC50 of 87.85 and 36.96 ppm in comparison to that of susceptible strain (53.61 and 19.89 ppm after 24 and 48 hrs, respectively). DISCUSSION From the results it is evident that both the field strains exhibit varying ratios of insecticide resistance against insecticides in the order of trichlorofon < bifenthrin < lambda-cyhalothrine < Malathion < spinosad. Both the field strains showed high degree of susceptibility towards spinosad. Hsu and Feng (2000) assayed B. dorsalis adults and found trichlorfon to be the least effective among the five insecticides tested. Our results are also in line to that of Kashyap and Hameed, 1982, but contradictory to the findings of Raga and Sato (2006) who reported trichlorofon as most effective against Ceratitis 166

RR 0.77 1.08 1.63 1.85

capitata. Slope and 2 value ( 1-2) revealed homogenous population of B. zonata being tested in the present assay. Resistance to malathion in similar species of fruit fly B. dorselis has been reported (Hsu et al., 2004) and present results of resistance to malathion also strengthen wide spread distribution of its resistance. Magana et al. (2007) have shown resistance to malathion in the field populations of Ceratitis capitata. The cross resistance pattern can also be witnessed in present study as populations were resistant to trichlorfon and malathion and also cyhalothrin and befintrhin, indicating two mechanism of resistance operating in these populations. Elevated hydrolytic and oxidative metabolic enzymes combined altered AchE with poor catalytic efficiency might contribute to resistance of B. dorsalis to malathion (Hsu et al., 2004). CONCLUSION Further studies on populations of fruit flies in different insecticides oriented farming systems will help to understand the biochemical mechanism of resistance. This study also suggests replacement of trichlorfon with spinosad. REFERENCES Abbott, S.W., 1925. A method of competing the effectiveness of an insecticide. J. Econ. Ent., 18: 265-267. Abdullah, K., M. Akram and A.A. Alizai, 2002. Nontraditional control of fruit flies in guava orchards in D. I. Khan. Pak. J. Agric. Res., 17: 195-96. Abdullah, K. and A. Latif, 2001. Studies on baits and dust formulations of insecticides against fruit fly (Diptera: Tephritidae) on melon (Cucumis melo) under semi arid conditions of D. I. Khan. Pak. J. Biol. Sci., 4: 334-335.

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