Assessments the toxic effects of entomopathogenic bacterium , Bacillus thuringeinsis subsp . kurstaki , and methomyl insecticide on larval instars of the greater sugarcane borer ; Sesamia cretica ( Lederer )

This study was conducted to evaluate the efficacy of the bio-insecticide Bacillus thuringiensis (Dipel 2x 6.4 % WP) against, 1 and 2 instar Larvae of Sesamia cretica compared with methomyl (Lannate 90 % SP). After 48 hours from treatment, The LC50 value of B. thuringiensis for 1 instar and 2 larvae were 0.00526 and 0.7 gm, respectively. While the LC50 of methomyl after 24h. for 1 instar and 2 larvae were 3394 and 5481 ppm, respectively. After treatment of 1 and 2 instars larvae of S. cretica with LC50 concentrates of B. thuringiensis, the average larval duration of S. cretica were (36.53 & 24.69) and (33.78& 20.31) days for treated and untreated, respectively. In addition, the pupation percentage was (47 & 92 %) and (18 & 84 %) for treated and untreated, respectively. The larval mortality percentage was (53 & 8 %) and (82 & 16 %) for treated and untreated, respectively. The pupal weight for (treated & untreated), S. cretica were (0.1887 & 0.184 gm) and (0.1842 & 0.1601 gm) when it treated as 1 and 2 instars larvae, respectively. Furthermore, the pupal duration were (12.14 & 10) and (10.44 & 9.83 days) for (treated & untreated), when S. cretica treated as 1 and 2 instar larvae, respectively. The total adult emergence for 1 instar and 2 instar larvae were (94 & 100%) and (100 & 100%) for treated and untreated, respectively. On the other hand results indicated that, after treatment of 1 and 2 instar larvae of S. cretica with LC50 concentrates of methomyl, the average larval duration of S. cretica were (31.41 & 24.69) and (33.87& 20.31) days for treated and control, respectively. The pupation percentage was (34 & 92 %) and (28& 84 %) for treated and untreated when S. cretica treated as 1 and 2 instars larvae, respectively. The larval mortality percentages were (66 & 8 %) and (72 & 16 %) for treated & untreated, respectively. The pupal weight for treated and control, were (0.1626 & 0.184 gm) and (0.1994 & 0.1601 gm), respectively. The pupal duration were (12.41 & 10) and (10.83 & 9.83) days for treated and untreated, respectively. The total emergence percentage for 1 instar larvae were (100 & 100 %) and (85.7& 100%) for 2 instar larvae for each (treated & untreated), respectively.


INTRODUCTION
The greater sugarcane borer; Sesamia cretica is considered the most serious corn bore in Egypt and attacks young maize plants shortly after emergence devours the whorl leaves and may kill the growing points, causing dead hearts.It is also capable of damaging older plants and excavating tunnels into the stems, ears and /or cobs.This pest lays its eggs during March, so its larvae cause complete death of small maize plants in April and May causing drastic yield losses (Mostafa, 1981;Simeada, 1985;El-Mitwally, 1987 andEl-Naggar, 1991).These losses are mainly attributed to the decrease in number of plants per unit area at harvest because of the large number of dead hearts, increase in plant lodging and eardrops and predisposing infested plants to disease organisms.
Over the years, it has been important for humans to control the populations of harmful insects and insecticides used for this purpose in agricultural and horticultural sectors.Synthetic insecticides, owing to their various side effects, have been widely replaced by biological insecticides.Bacillus thuringiensis (BT) accounts for 90 % of the bioinsecticide market and it produces insecticidal toxins referred to as delta endotoxins (Chattopadhyay et al. (2004).The aim of the present work is to evaluate the toxicological activity and latent effects on some biological aspects of the entomopathogenic bacterium, Bacillus thuringeinsis subsp.kurstaki, and chemical insecticide methomyl against 1 st and 2 nd instar larvae of the greater sugarcane borer; Sesamia cretica (Lederer).

MATERIALS AND METHODS
The present study was carried out to study the effect of Bacillus thuringiensis and methomyl insecticide against 1 st and 2 nd instar larvae of the greater sugarcane borer; Sesamia cretica (Lederer) under laboratory conditions.

Test insect
Infested corn plants with S. cretica (larval stage) were taken from the field and transferred to the laboratory.The larvae of S. cretica were reared on maize pieces at a controlled conditions in electric incubator at 27˚C ± 2 and relative humidity of 65% ± 10 % R. H., as follows: 1-Obtained larvae kept, individually, in plastic cups containing a layer of moistened sawdust on the bottom (1.5cm thickness).Every two days, larval feces removed as well as dried stems, which replaced by fresh maize stems or young corn ears, always in sufficient amounts.Larvae left under laboratory conditions at 27˚C ± 2 and relative humidity of 65% ± 10 % R. H. until pupation.2-The obtained pupae were sexed and introduced singly into 1 x 3" glass vials, each provided with a small piece of moistened cotton wool at its bottom, and plugged with cotton wool.3-Vials were kept under the same laboratory conditions mentioned above until adult emergence.4-Immediately after emergence, every couple placed in a glass chimney cage fixed on a plastic cup filled with fine sand covered on the top by muslin cloth kept in position by a rubber band.Inside each cage, 4 wax papers were rolled and fixed in the sand to act as an oviposition site for S. cretica moths.5-Pairs replicated in 10 times and each jar covered with pin notched wax paper to provide sufficient ventilation.6-The daily-obtained egg-masses kept in Petri-dishes containing small pieces of moistened cotton wool, where eggs were counted and left until hatching.7-Each 10 newly hatched larvae were introduced together into a glass jar containing 10 small pieces (about 10 cm long) of the fresh tender rolled leaves surrounding the growing point of maize plants.
8-Jars tightly covered with muslin and inspected every two days to renew maize leaves.9-Immediately after the second larval instar, small cutting (about 10 cm long) of fresh maize plant stems or young corn ears were used for feeding.Cuttings were renewed every two days until pupation.

Bioassay studies
Laboratory experiments were conducted to study the toxic effects of Bacillus thuringiensis (Dipel 2X) as a biocontrol agent and Methomyl (Lannate) as a carbamate insecticide on 1 st , and 2 nd instars of S. cretica larvae.For treatment, three equal pieces of tender parts stems of maize plant dipped in the desired solution for about two minutes after which the treated parts were left in shade for about 10 minutes to dry.The experimented larvae kept starved for about 4 hours, before offering the treated food to assure rapid ingestion.Larvae were offered contaminated maize parts for 48 h.when the tested compound is B.t. and for 24 when the tested compound is methomyl.The total number of treated larvae / treatment was 50 that divided in 5 replicates of 10 larvae each.The same numbers of larvae considered as a control, these larvae offered tender parts of maize plant stems immersed in distilled water.Results were presented graphically as log / probity regression lines and LC 50 were values calculated by the computer program Sigma Plots for Windows (version2).Furthermore, LC 50 of tested compound were calculated for both the 1 st and 2 nd instars larvae.Fresh tender parts of maize plant stems immersed in the LC 50 of each tested compounds and then left to dry at room temperature before contaminated maize stems as before for each compound.Each treatment comprised 50 larvae and replicated 5 times (larvae/jar).The same numbers of larval mortality was determined at the end of the larval stage.Corrected mortality counts according to formula (Abbott, 1925), then submitted to probit analysis using Finney (1971) thuringiensis increased by increasing of used concentration, the corrected mortality % were 82.2, 60.0, 53.3, and 24.4 % when the B. thuringiensis concentrations were 0.0625, 0.015625, 0.0039 and 0.00097 ppm, respectively.On the other hand, the LC 50 of B. thuringiensis for 1 st instar larvae was 0.00526 ppm.The slope for S. cretica 1 st instar larvae when treated by B. thuringiensis was 0.8323.Results also, indicated that the toxic effect of B. thuringiensis increased by increasing the concentrates, the mortality % were 72.92, 62.5, 50, 41.67, 18.75, 14.5, 6.25  2-Toxic effect of methomyl against the 1 st and 2 nd instars larvae of S. cretica Results in Table (2) show the toxic effect of methomyl increased by increasing of used concentration, the corrected mortality percentage were 91.56, 73.33, 42.22, 26.67, 22.22and 2.22 % methomyl concentrations were 14062.5, 7031.25, 3515.6, 1757.8, 878.9 and 439.5 ppm, respectively.On the other hand, the LC 50 of methomyl for 1 st instar larvae was 3394 ppm.The slope for 1 st instars larvae of S. cretica when treated by methomyl was 2.0428.Ebieda et al. (1998)   Results in Table (2) indicated that the toxic effect of methomyl increased by increasing of used concentration, the mortality % were 91.67, 60.42, 18.75 and 10.42 % when the methomyl concentrations were 14062.5, 7031.25, 3515.6 and 1757.8ppm, respectively.Where as the LC 50 of methomyl against 2 nd instar larvae was 5481 ppm.The slope for 2 nd instar larvae of S. cretica when treated by methomyl was 3.0148.

3-Toxicity of B. thuringiensis and methomyl on some biological aspects of S. cretica
This part of study aims to investigate the latent effect of LC 50 of tested compounds on certain biological aspects; larval duration, pupation, larval mortality, pupal weight, pupal duration, pupal mortality and emergence percentage of the S. cretica.Such investigations may throw a light to complete the picture on the mode of actions of B. thuringiensis and methomyl during the 1 st generation.

1-On 1 st instars larvae
Data in Table (3) showed that, after treatment the 1 st instar larvae of S. cretica with LC 50s of B. thuringiensis and methomyl the average larval duration of S. cretica were 36.53,31.42 and 24.69 days for B.t, methomyl and untreated, respectively.The pupation percentage was 47.0, 34.0 and 92.0 for B.t, methomyl and untreated, respectively.The larval mortality percentage was (53.0, 66.0 and 8.0).
Data in Table (3) indicated that, the pupal weight for treated and untreated S. cretica was 0.1887, 0.1626 and 0.184 gm for B. thuringiensis, methomyl and untreated, respectively.When it treated as 1 st instars larvae, respectively.On the other hand, the pupal duration was 12.14, 12.41 and 10 days for B. thuringiensis, methomyl and untreated, respectively.For pupal mortality percentage, it was 6.67, 0.0 and 0.0 for B. thuringiensis, methomyl and untreated, respectively.thuringiensis, methomyl and untreated, respectively.For pupal mortality percentage, it was 0.0, 14.3 and 0.0 for B. thuringiensis, methomyl and untreated, respectively.
El-Halim (1993) evaluated the insecticidal activity and the latent effect of Dipel 2X, a commercial preparation of B. thuringiensis subsp.kurstaki in the laboratory on S. littoralis larvae.Dipel 2X had slight insecticidal activity against the cotton leafworm.The ability of larvae to recover decreased with the increase in concentration and/or feeding time.Both larval and pupal duration markedly prolonged with dose increase, while percentage pupation reduced.Marked latent adverse effects detected on adult emergence, fecundity and egg viability, particularly with doses above 320 IU/ml.Assessments the toxic effects of entomopathogenic bacterium, Bacillus thuringeinsis 7 Data in Table (5) and Figs.(1 and 2) showed the total emergence %, sex ratio % and malformed pupa and adult for 1 st and 2 nd instars larvae were 94, 100, 100 100.0, 85.7and 100.0 for B.t, methomyl and untreated, respectively.

Fig 1 :
Fig 1: Malformed adult of S. cretica following treatment with methomyl as 1 st instar

-Toxic effect of B. thuringiensis against the 1 st and 2 nd instars larvae of S. cretica
Data summarized in Table (1)showed that the toxic effect of B.

Table 1 :
Toxicity of different concentrations of B. thuringiensis against, 1 st and 2 nd instars larvae of S. cretica.

Table 2 :
Toxicity of different concentrations of methomyl against, 1 st and 2 nd instars larvae of S. cretica.

Table 3 :
Effects of B. thuringiensis and methomyl on larval and pupal stages of S. cretica after treated as 1 st instar larvae with LC 50 concentrations.

Table 4 :
Effects of B. thuringiensis and methomyl on larval stage of S. cretica after treated as 2 nd instar larvae with LC 50 concentrations.The values have the same letters horizontally are non-significant different.

Table 5 :
Effects of B. thuringiensis and methomyl on adult stage of S. cretica after treated as 1 st instar and 2 nd instar larvae with LC 50 concentration