Baseline Susceptibility of Diamondback Moth Plutella xylostella L (Lepidoptera:Plutellidae) to chlorantraniliprole 18.5 SC in Tamil Nadu

The baseline susceptibility of diamondback moth, Plutella xylostella L. to chlorantraniliprole 18.5 SC was studied in the Insecticide

field. To date, the pest has developed resistance to 81 insecticides (Anonymous, 2009) and has become one of the most difficult pests to control in cruciferous vegetables.
Chlorantraniliprole is the first pesticide from the anthranilic diamides and it was found to provide broad spectrum activity within Lepidoptera (Temple et al., 2009). It acts as selective agonist for ryanodine receptors in Lepidopteran insects. It causes unregulated Ca 2 + release from intracellular calcium stores, which results in the insect's ability to regulate muscle function being impaired, generating poisoning symptoms that include: rapid feeding cessation, lethargy, muscle paralysis, and ultimately insect death (Lahm et al., 2005;Sattelle et al., 2008). It was relatively harmless to beneficial arthropods and has not been found to exhibit cross resistance with existing insecticides (Lahm et al., 2009). These favourable characteristics was found useful as an additional management tool to control P.xylostella and a good fit for integrated pest management (IPM) since its introduction in India during 2008.
However, repeated applications of chlorantranili prole at higher dose on cauliflower and cabbage against P.xylostella might result in resistance development in the high intensity production area. The present study was undertaken to elucidate information on baseline susceptibility of P. xylostella to chlorantraniliprole 18.5 SC and resistance status of chlorantraniliprole in cauliflower and cabbage growing areas of the Tamil Nadu.

Materials and Methods
Laboratory studies were carried out in the Insecticide Resistance Laboratory, Department of Agricultural Entomology, Tamil Nadu Agricultural University, Coimbatore. The larvae collected from cauliflower and cabbage growing areas of Coimbatore (Thondamuttur) was reared up to 25 generations without exposure of insecticides using cauliflower leaves maintained in pot culture. Mass culturing of P. xylosella was carried out by following the method described by Liu and Sun (1984) with little modifications. Mustard seedlings were used for egg collection in oviposition cage and same cage was used for larval rearing. Raising of mustard seedling and rearing of P.xylostella was done under the lab conditions at 12:12 (L:D) and at prevailing room temperature 28 ± 2 º C . The third instar larvae (0.5±0.1 cm; 1.75±0.25 mg) were collected on tenth day after egg laying for conducting bioassay.
The dilutions required for bioassay were prepared from the formulated product of known purity of insecticide using distilled water. The insecticide formulation used was chlorantraniliprole 18.5 SC (Coragen 18.5 SC M/s E.I. DuPont India Pvt. Ltd, Gurgaon, Haryana.) The leaf disc bioassay method originally described by Hirano (1979) further supplemented by Tabashnik and Cushing (1987) was adopted. The leaf discs of 6 cm diameter were cut covering either side of the midrib. These leaf discs were dipped in test concentrations for about a min, and then removed; excess fluid drained and air dried by hanging to a thread for about one hour and transferred to a small round plastic container of 7 cm height and 6 cm dia. over a moistened filter paper. The leaf discs were placed slantingly to rest on the sides of container so that larvae can move on either side. Ten larvae, starved for 12 h, were transferred to each disc and the container was covered with lid provided with holes for aeration which constituted one replication. Each treatment was replicated three times and observations on mortality of the larvae were taken at 24h after treatment.
For generation of baseline data, the insects were reared without exposure to insecticides and cultured continuously without selection pressure throughout F 1 to F 25 generations. Bioassays were conducted to construct LCPM lines for a susceptible population. Based on lethal concentrations obtained for the test insecticide a tentative discriminating dose was fixed based on LC 95 value of the F 25 generation. The susceptible indices(SI), rate of resistance decline (R) and the number of generations required for tenfold decrease in the LC 50 value (G) were calculated (Regupathy and Dhamu, 2001).
Monitoring studies was done at weekly interval from January 2013 to March 2013, in major cauliflower and cabbage growing areas of Tamil Nadu viz., Coimbatore, Oddanchatram and Udaghamandalam. Frequently used insecticides in cauliflower and cabbage viz., chlorantraniliprole 18.5 SC (Coragen®), spinosad 2.5 SC (Success®), emamectin benzoate 5 SG (Proclaim®), and quinalphos 25 EC (Ekalux®) were selected for resistance monitoring. Test insecticides were used at discriminating concentrations viz., chloran traniliprole 23 ppm (obtained from base line data) spinosad 12 ppm, emamectin benzoate 2 ppm (Lavanya et al., 2010) and quinalphos 3 ppm (Chandrasekaran and Regupathy, 1996). The larvae collected at weekly intervals were reared upto F 1 generation and third instar (1.83±0.28 mg) larvae were used for the bioassay. Leaf disc method was followed for bioassay with each treatment comprising of 60 larvae. The mortality was recorded at 24 h after treatment and resistance percentage was calculated by using Abbott's formula (Abbott, 1925).

Results and Discussion
The LC 50 and LC 95 values of chlorantraniliprole 18.5 SC for 25 generations against P.xylostella are presented in Table 1. The LC 50 for F 1 population was 20.06 ppm and LC 95 was 835.68 ppm. The susceptibility was found increasing over generations. In terms of LC 50, susceptibility increased up to F 22 generation and got stabilized from F 23 to F 25 generations (0.91 ppm). The susceptibility in terms of LC 95 also increased up to F 23 generation (23.26 ppm) and got stabilised in F 24 and F 25 generations (23.11 ppm).
The susceptibility indices based on LC 50 and LC 95 were 22.02 and 36.15, respectively after F 25 generations. (Table.2). The number of generations required for 10 fold decrease in LC 50 was 20 generations. The rate of resistance decline (R ) was -0.0505. The computed LC 50 and LC 95 values indicated that the susceptibility gradually increased with succeeding generations from F 1 to F 25 (20.06 ppm to 0.91ppm) and similarly LC 95 values from F 1 to F 25 decreased from 835.68 ppm to 23.11 ppm. A tentative discriminating dose (DD) 23 ppm was arrived based on LC 95 value of 23.11 ppm computed for F 25 generation and it recorded 99-100 per cent mortality of the susceptible population. Similar studies made by Wang et al. (2010) reported that the variation among 16 field populations was low (fivefold) with chlorantraniliprole and LC 50 values ranged from 0.221 to 1.104 mg/litre. The discriminating dose of 23 ppm computed in the     Wang and Wu (2012), who stated that six field populations from southern China (Guangdong Province) showed higher LC 50 values 2.6, 12, 18, 81, 140, and 2,000 fold than LC 50 of Roth (Susceptible strain), which would be due to intensive use and misuse of chlorantraniliprole. The present line of work is also in agreement with Lavanya et al.(2010), who had reported that emamectin benzoate (2 ppm) and spinosad (12 ppm) recorded 5.81 to 15.6 and 34 to 15.73 per cent resistance with all the three diamondback moth populations of Coimbatore, Ooty and Oddanchatram. Similarly Zhao et.al.(2006) reported that, population collected from collards from Oxnard CA showed low level of resistance to spinosad in 2002 (TR, 27.3 fold with 20% survival), but it increased rapidly during 2004 (Toxicity Ratio >15,000 fold and 93% survival). Patil (2011) reported that LC 50 of emamectin benzoate was in the range of 9.16 to 39.07 ppm with maximum in Naski strain and minimum in Wadegoan (Akola) indicating that emamectin benzoate showed higher degree of resistance. Present findings from monitoring studies indicate that resistance in P.xylostella to chlorantraniliprole is in the initial stage, whereas high levels of resistance to spinosad and emamectin benozoate recorded might due to long time and repeated use in the test regions.