EFFECT OF DIFFERENT AGAINST , INDICOLA L. (ISOPTERA: TERMITIDAE) AS SLOW ACTING TOXICANTS

AHMAD-UR-RAHMAN SALJOQI1*, NOOR MUHAMMAD1, IMTIAZ ALI KHAN2, SADUR-REHMAN3, MUHAMMAD NADEEM2and MUHAMMAD SALIM1

1Department of Plant Protection, The University of Agriculture, Peshawar- 2Department of Entomology, The University of Agriculture, Peshawar-Pakistan 3Agriculture Research Institute, Tarnab, Peshawar, Pakistan. *Corresponding author: [email protected]

ABSTRACT Three insecticides viz Regent, Tracer and Match were evaluated as slow toxicants against subterranean termites, L (Isoptera:Termitidae). The experiment was performed using workers of H. indicola to which these insecticides were dipped in blotting paper once only, in start with the following concentration i.e. 0.000312, 0.000156, 0.00078, 0.00039 and 0.000195%. The total mean percent of mortality after ten days results concluded that Regent ( 5% SC) was 91.79, 86.40, 78.94, 74.18 and 62.06%, while Tracer (Spinosad 240 SC) was 72.60, 63.53, 60.60, 59.06 and 28.60%, and finally Match (Lufenuron 5% EC) was 49.40, 31.06, 26.19, 22.18 and 10.66%, repectively. Maximum mortality and avoidance was obtained by toxicity of Regent, as it was capable of obtaining 100% mortality even before day ten on every concentration therefore, Regent was considered highly toxic. Tracer was found to be slow acting agent. 100% mortality was recorded by using the 1st concentration of Tracer on day 7, on day 9 by using the 2nd concentration, while on day 10 by using the 3rd concentration of Regent. Match was unable to cause 100% mortality by using any of the tested concentrations. Based on the mortality recorded among the three insecticides the last two (0.000039 and 0.0000195%) concentrations of Regent caused 100% mortality on the 7th and 8th days. The 1st three (0.000312, 0.000156 and 0.000078%) concentrations of Tracer caused 100% mortality on the 7th, 8th and 9th day, and the 1st concentration of Match caused 80% mortality on the 10th day. These concentrations were found the most effective and are recommended as slow acting toxicant in the baits for H. indicola control. Keywords: Insecticides, different insecticides concentrations, Heterotermes indicola, Slow acting toxicants, Termites mortality Citation: Saljoqi. A. U. R., N. Muhammad., I. A. Khan., M. Nadeem., S. Rehman and M. Salim. 2014. Effect of different insecticides against termites, heterotermes indicola l. (isoptera: termitidae) as slow acting toxicants. Sarhad J. Agric. 30(3): 333-339

INTRODUCTION Termites are widely distributed throughout Pakistan, especially in Khyber Pakhtunkhwa region. They are either soil or wood inhibiting termites. The climate of Khyber Pakhtunkhwa is most favorable for production of different fruits, food and sugar crops. Among fruits, peach, apricot, citrus, guava, plum, apple and persimmon are widely grown and are attacked by different pests, but termite is the most serious one, which inflicts substantial losses to farming community (Salihah et al., 1994). The termites, Heterotermes, Coptotermes and Odontotermes have been observed to infest apricot, pear, plum, peach and lemon causing different ranges of infestations (Salihah et al., 1992) Sugarcane, Saccharum officinarum is an important cash crop of Pakistan. During 1993-1996, the reduction in area and production was due to the attack of a number of but the termite was on the top. In 1985 complaints were seen in different newspapers that sugarcane crop has been destroyed by termites in different areas of Peshawar District (Salihah et al., 1998). Besides crops and buildings, fruit trees are also damaged by Heterotermes indicola L. Successful methods of control have not been developed largely due to the lack of information on organization of termite colonies, which do not build mounds or any other readily detectable structures (Spragg and Fox, 1994). All the 15 species of termites infesting agricultural crops recorded in NWFP are mound less and have no detectable structures (Salihah et al., 1994). Highly effective chemical treatments have been available for many years to prevent subterranean termites' attack and to control infestation. The frequent use of fast-acting termiticides for control of termites has generated a number of Ahmad-ur-Rahman Saljoqi, et al. Effect of different insecticides against termites… 334 biological and environmental hazards in air, water, soil and food. Furthermore, these treatments are expensive and require specialized techniques. Conventional can kill beneficial insects (parasitoids and predators), which feed upon insect pests. Just one spray can upset the balance between pests and the beneficial predators (www.paceproject.net). Artificial chemicals can stay in the environment and in the bodies of causing problems for many years. Pests become resistant to pesticides so more powerful chemicals are needed (Parihar and Singh, 1992). Kamdem et al. (2002) used Fipronil protection against decay fungi and termites. Fipronil, an already in use to protect agricultural crops and used in the public health sector, is added in the formulation of phenol- formaldehyde resin during the laboratory manufacture of 5-ply plywood. Fipronil loading is sufficient to control the termite activity on plywood samples (Smith and Rust, 2007). Subterranean termites are serious pests of fruit trees, food crops and buildings in Pakistan, especially in Khyber Pakhtunkhwa. Termites commonly known as ‘white ’ inhibit nests of various kinds. They have well developed caste systems, which include king, queen, soldier, worker and nymphs. Control of these termites has generally been erratic and unsatisfactory because of their cryptic nature, long life, huge colony size and extensive range of foraging territories (Salihah et al., 1994). Traditionally soil treatments have been conducted where insecticides of high repellency and long residual effect have been used for flooding the target area. This method is costly, inefficient and environmentally unsafe (Su et al., 1993). Use of slow acting toxicant on the other hand is a safe method of managing termite populations. These slow acting toxicants should be palatable so that the termites foraging carry these chemicals back to the colony and distribute among their nest mates (Sattar, 2002). For the control of this destructive pest a number of chlorinated hydrocarbons like Aldrin, Dieldrin, Chlordane, Heptachlor, DDT, and BHC are known effective against termites. Environmental Protection Agency (EPA) bans their use in the world because of environmental hazards they cause. Keeping in view the economic importance of H. indicola as a common problematic pest of agricultural crops and buildings, laboratory trials were conducted to find out the effect of different insecticides for its management as a slow acting toxicant (a slow acting insecticide is one that causes less than 15% mortality after 24 hours of exposure and more than 84% mortality after 14 days).

MATERIALS AND METHODS This study was conducted to evaluate the effects of different insecticides against H. indicola as slow acting toxicants at Entomology Division of Nuclear Institute for Food and Agriculture (NIFA), Tarnab, Peshawar, during September, 2008 to November, 2009. H. indicola were collected from the infested buildings around the NIFA by using NIFA– TERMAP technique (Salihah et al., 1998). The different insecticides used in this experiment are given below.

Table 1. Different insecticides used in the experiment S. No. Chemical Name Trade Name Groups Mode of action Dose ha-1 1. Fipronil (5% SC) Regent Pyrazoles Chitin blocker 1250 ml 2. Spinosad (240 SC) Tracer Spinosyns Stomach Poison 750 ml 3. Lufenuron (5% EC) Match Cyromizine Growth Regulator 500 ml 4. Control Distilled Water

Detection of Termites The foraging points of termites were detected by using stake method as well as visually observing termites' galleries. Wooden stakes (2.5x4.0x28 cm) made of poplar were driven 25 cm deep into the soil in grid pattern (2.4x2.4 m) around building and in orchard and was observed fortnightly. Active foraging points were marked and infested stakes were replaced by NIFA-TERMAP. Collection of Termites Termites were collected fortnightly. Infested bundles of NIFA-TERMAPs were replaced with new bundles. Bundles were made from five slices of popular wood blotting paper and rubber band round them. The infested traps were brought to the laboratory. Termite along with the soil and debris was passed through different mesh sieves placed one above the other in a plastic tube of 29.5 cm in diameter to separate the termites from the soil and debris. A pair of moist blotting paper was placed to separate soil and termites for few minutes, hereafter, thousands of termites were Sarhad J. Agric. Vol.30, No.3, 2014 335 then collected from the blotting paper. Then the cleaned termite-workers and -soldiers along with a pair of moist blotting paper was placed in the Petri-dishes and kept in desiccators for moisturizing of termites. Once termites were collected, insecticides were prepared in five different concentrations. All these concentrations were tested and compared with the control. For each concentration, a total 30 Petri-dishes were prepared by placing a double layer of blotting paper and the blotting paper was soaked with insecticides by dipping it into the solution until it became completely wet. Fifty termite workers were kept in each Petri-dish and the dishes were kept in the desiccators for a period of 10 days. The data was recorded on a daily basis taking by randomly from each Petri-dish. For each insecticide testing along with control experiment 180 Petri-dishes were used. Data collected right from the next day of the beginning of experiment on mortality, and then those Petri-dishes were discarded. This process was repeated for a period of 10 days. The experiment was conducted under controlled conditions of 28±2°С and 60±5% R.H. The experiment was laid out as Two-factorial in a Randomized Complete Block Design (RCBD). Preparation of Toxicant Bait Three insecticides; Regent, Tracer and Match were selected to be tested as slow acting toxicants. Prior experiments, fresh concentrations of the tested pesticides were prepared by diluting in distilled water. Five concentrations (0.000312, 000156, 0.000078, 000039 and 0.0000195%) of each insecticide were tested against H. indicola. All these concentrations were compared with the control (distill water). A pair of blotting paper was soaked in each concentration and was placed in each Petri-dish for a period of 10 days along with termites' workers. Statistical Analysis All the collected data were analyzed by using MSTATC computer package and the means were separated by using Duncan’s Multiple Range Test with 5% probability level (Steels and Torrie, 1960).

RESULTS AND DISCUSSION The toxicity results of applying Reagent, Tracer and Match in five different concentrations against termites evaluated as slow acting toxicant for a period of 10 days under laboratory condition are presented in Tables 2-4 and Fig. 1-3. Table 2 shows mortality of H indicola by using Regent (Fipronil-5% SC) applied at 0.000312, 0.000156, 0.000078, 0.000039 and 0.0000195% concentrations for a period of ten days. The Table shows that all concentrations were found very effective against termites as compared with control. Maximum percentage of termite mortality was recorded on the highest concentration (0.000312%) and the mortality was found on decreasing rate as the concentrations of the Regent decreased. The result showed that the first three higher concentrations showed termites' mortality in a short time according to their dosage rate, while the last two lowest concentrations took long time for getting the 100% mortality, which were found perfect as slow acting toxicants.

Table 2. Effect of different concentrations of Regent (Fipronil-5%SC) against H eterotermes indicola for 10 days after treatments under controlled laboratory conditions. Days Concentrations (%) 0.000312 0.000156 0.000078 0.000039 0.0000195 Control 1 25.33 c 8.66 d 6.00 e 3.33 f 1.33 f 0.00 g 2 92.60 b 69.70 c 35.00 d 26.16 e 17.43 e 0.66 g 3 100.00 a 89.63 b 66.16 c 49.26 d 21.53 d 4.00 g 4 100.00 a 100.00 a 85.10 b 75.90 c 31.90 d 6.00 ef 5 100.00 a 100.00 a 97.13 a 87.16 b 71.16 c 7.33 e 6 100.00 a 100.00 a 100.00 a 98.00 a 81.03 c 8.66 de 7 100.00 a 100.00 a 100.00 a 100.00 a 96.30 b 10.66 cd 8 100.00 a 100.00 a 100.00 a 100.00 a 100.00 a 13.33 bc 9 100.00 a 100.00 a 100.00 a 100.00 a 100.00 a 16.00 ab 10 100.00 a 100.00 a 100.00 a 100.00 a 100.00 a 17.33 a Means followed by the same letters are not significantly different at 5% level of probability (DMR-test). Fig. 1 shows the total mean percent mortality of termites by using different concentrations of Regent for a period of 10 days. All concentrations were found effective as compared with control. Maximum total mean percent mortality of termites (91.79%) was obtained by using the highest concentration (0.000312%). Decreasing mortality rate of termites was recorded as the concentrations were decreased. The lowest total mean percent mortality of termites (62.06%) was recorded by using the lowest concentration (0.000195%). Ahmad-ur-Rahman Saljoqi, et al. Effect of different insecticides against termites… 336

100 91.79 a 86.79 b 90 78.93 c 80 73.98 d

70 62.06 e

60

50

40

30

20 Total Mean Percent Mortality 8.37 f 10

0 0.000312% 0.000156% 0.00078% 0.00039% 0.000195% Control Concentrations (Treatments)

Fig. 1 The effect of different concentrations of Regent-(Fipronil-5%SC) on the total mean percent mortality Heterotermes indicola for a period of 10 days under controlled laboratory conditions. Different small letters above bars indicate significant differences at 5% level of probability (DMR-test).

Table 3 showed the mortality of H. indicola by using Tracer (Spinosad- 240 SC). It shows that all concentrations were found very effective against termites as compared with control. Maximum percentage of termite mortality was recorded for the highest concentration. Mortality was found on decreasing rate as the concentration of the Tracer decreased. The 1st concentration (0.000312%) caused 9.33, 16.66, 37.33, 70.00, 94.00, 98.66 and 100% mortality on day 1st to day 7th, respectively, and the mortality increased gradually day by day. At the same time, the second concentration (0.000156%) caused mortality of 8.66, 14.33, 30.00, 41.33, 73.33, 86.66, 88.66, 94.33 and 100% mortality on day 1st to day 8th, respectively. If we compare it to other concentrations and control it was much better because the mortality rate was slightly low from other concentrations and high from control. The third concentration (0.000078%) was used for the same time and it caused 6.00, 11.33, 22.66, 40.00, 69.33, 82.33, 84.33, 91.00 and 100% mortality from day 1st to day 9th, respectively. The 3rd concentration also gave good results because both 2nd and 3rd concentrations were acceptable for slow acting toxicant. The concentration of 0.000039% caused mortality of 5.33% on first day, followed by 10.66% on day 2nd, 32.66% on day 3rd, 39.33%on day 4th, 62.00% on day 5th, 70%on day 6th, day 80% on 7th and 82.33, 91.33 and 98% on day 8th, 9th and 10th, respectively. Last and low concentration used was 0.0000195%, and caused 3.66%-48.66% mortality from day 1st up to day 10th. The results showed that the higher concentrations (1 to 3) showed 100% mortality to termites at a time according to their definition, while the last two lowest concentrations took long time for getting the 100% mortality, which was not required for slow acting toxicants.

Table 3. Effect of different concentrations of Tracer (Spinosad-240 SC) against Heterotermes indicola for 10 days under controlled laboratory conditions. Days Concentrations (%) 0.000312 0.000156 0.000078 0.000039 0.0000195 Control 1 9.33 d 8.66 ef 6.00 f 5.33 g 3.66 h 0.00 e 2 16.66 d 14.33 e 11.33 ef 10.66 f 8.00 gh 0.00 e 3 37.33 c 30.00 de 22.66 e 32.66 ef 15.33 fg 4.66 d 4 70.00 b 41.33 d 40.00 d 39.33 de 20.00 ef 7.33 cd 5 94.00 a 73.33 c 69.33 c 62.00 cd 27.33 de 10.66 bc 6 98.66 a 86.66 bc 82.33 bc 70.00 bc 30.66 cd 12.00 b 7 100.00 a 90.66 a 84.33 b 80.00 abc 36.66 bc 12.00 b 8 100.00 a 100.00 a 91.00 a 82.33 ab 43.33 ab 13.33 b 9 100.00 a 100.00 a 100.00 a 91.33 a 47.33 a 18.00 a 10 100.00 a 100.00 a 100.00 a 98.00 a 48.66 a 20.00 a Means followed by the same letters are not significantly different at 5% level of probability (DMR-test). Sarhad J. Agric. Vol.30, No.3, 2014 337

Fig. 2 shows the total mean percent mortality of termites by using different concentrations of Tracer for a period of 10 days. All concentrations were found effective as compared with control. Maximum total mean percent mortality of termites (72.60%) was obtained by using the highest concentration. Decreasing mortality rate of termites was recorded as the concentrations were decreased. The lowest total mean percent mortality (28.09%) of termites was recorded by using the lowest concentration 0.000195%. The Table 4 shows the mortality of termites by using Match (Lufenronu-5% EC). Concentrations used were 0.000312, 0.000156, 0.000078, 0.000039 and 0.0000195%. The Table shows that all concentrations were found very effective against termites as compared with control where none of the insecticides were applied. Maximum percentage of termite mortality was recorded for the highest concentration, then the mortality was found on decreasing rate as the concentrations of the Match decreased. The 1st concentration caused mortality of 13.33%-78.00% from the day 1st to day 10th, much better results from the last four concentrations and control. It was enough for the slow acting toxicant. The 2nd concentration caused 8.66% mortality on day 1st and 54.66% up to day 10th, so, it did not cause 100% mortality up to day 10th, therefore, the mortality rate was too low. The 3rd concentration was used in the same manner and it caused 6.66% mortality on day 1st up to 48.00% on day 10th, but the mortality rate was very low so it could not reach to 50% mortality. It was very low mortality rate, therefore, not suitable for slow acting toxicants. According to definition, if the mortality started before 24 h and after 72 h, thus, it would be not considered as a slow acting toxicant. The 4th concentration caused mortality from day 1st to day 10th as 5.33%-42.66% and also very low results. The 5th concentration caused mortality from day one to day ten as 0.00-25.33%. The last concentration was too low from required rate, where the results of 5th concentration and control were the same; therefore, it couldn’t keep in the bait blocks for termites control. The result showed that the 1st concentration gave 100% mortality of termites in a time according to their definition, while the last four lowest concentrations took long time for getting the 100% mortality, which was not suitable for a slow acting toxicant.

100 72.58 a 90 63.73 b 80 60.69 b 70 52.16 b 60 50 40 28.09 c 30 20 9.79 d 10

Total Mean Total Percent Mortality 0 0.000312% 0.000156% 0.00078% 0.00039% 0.000195% Control

Concentrations (Treatments)

Fig. 2 The effect of different concentrations of Tracer (Spinosad- 240 SC) on the total mean percent mortality of H eterotermes indicola for a period of 10 days under controlled laboratory conditions. Different small letters above bars indicate significant differences at 5% level of probability (DMR-test).

Figure 3 shows the total mean percent mortality of termites by different concentrations of Match for a period of 10 days. All concentrations were found effective as compared to control. Maximum total mean percent mortality (49.39%) of termites was obtained by using the highest concentration (0.000312%). Decreasing mortality rate of termites was recorded as the concentrations were decreased. The lowest total mean percent mortality of termites (10.66%) was recorded by using the lowest concentration (0.000195%). Ahmad-ur-Rahman Saljoqi, et al. Effect of different insecticides against termites… 338

100 90 80 70 49.39 a 60 50 30.66 b 26.91 bc 40 22.16 c 30 10.66 d 20 7.53 e

TotalMeanMortalityPercent 10 0 0.000312% 0.000156% 0.00078% 0.00039% 0.000195% Control Concentrations (Treatments)

Fig. 3 The effect of different concentrations of Match (Lufenronu-5% EC) on the total mean percent mortality of Heterotermes indicola for a period of 10 days under controlled laboratory condition. Different small letters above bars indicate significant differences at 5% level of probability (DMR-test).

Regent, (Fipronil-5%SC) Chitin blocker systematically kills termites both with contact and stomach activity. Regent is used for the control of many soil and foliar insects on a variety of crops. It is also formulated as baits for cockroaches, ants and termites. Fipronil is effective against insects resistant or tolerant to pyrethrooid, organophosphate and carbamate insecticides. Fipronil blocks the gamma aminobutric acid (GABA) regulated chloride channel in nervous system, thus antagonizing the caiming effects of GABA similar to the action of the cyclodienes. In the present study the lower concentration of toxicity of Regent 0.00039% and 0.000195% caused 100% mortality on day seven and day eight which was enough found to act as slow acting toxicant but the higher concentrations were found too high, because they caused 100% mortality in a very short time. Toxicity was very high that is why it killed termite workers on spot. It did not give time to termites to go back to the colony and feed their nest mates. Regent gave high percentage of mortality, therefore it cannot be recommended as a slow acting termicides. The results of the present study are consistent with that of Manzoor et al. (2012), who found that Regent- Fipronil-5%SC effectiveness was very high at the higher concentrations, while they also found it acts as a slow acting toxicant at lower concentrations.

Table 4. Effect of different concentrations used against H eterotermes indicola of Match (Lufenronu-5% EC) for 10 days under controlled laboratory conditions. Days Concentrations (%) 0.000312 0.000156 0.000078 0.000039 0.0000195 Control 1 13.33 c 8.66 i 6.66 g 5.33 e 0.00 e 0.00 e 2 16.00 c 15.33 h 12.66 f 8.66 e 1.33 de 0.00 e 3 20.00 c 18.00 gh 16.00 ef 10.00 de 4.66 de 0.00 e 4 46.66 b 24.66 g 18.00 e 12.00 de 4.66 de 1.33 e 5 54.00 b 28.66 ef 22.66 d 16.00 de 5.33 d 2.66 de 6 62.00 ab 32.00 de 26.66 c 20.00 d 10.00 cd 5.33 cd 7 64.00 ab 36.00 cd 32.66 b 30.33 c 15.33 bc 6.00 c 8 66.66 ab 41.33 c 38.00 b 37.33 b 18.00 abc 8.00 bc 9 73.33 a 47.33 bc 40.66 b 39.33 ab 22.00 ab 9.33 b 10 78.00 a 54.66 a 48.00 a 42.66 a 25.33 a 12.66 a

Means followed by the same letters are not significantly different at 5% level of probability (DMR-test). Tracer, (Spinosad –240 SC) acts as a contact and stomach poison. Spinosad is a fermentation metabolite of the soil- inhabiting actinomycete, sacharopolyspora spinosa, . It has a novel molecular structure and mode of action that provides excellent crop protection typically associated with synthetic insecticides. In the present study the second, third and fourth concentrations were found desirable, because they caused the moderate mortality. The average percent mortality, 72.60, 63.53, 60.60, 59.06 and 28.60% was recorded for a period of ten days from 1st to 5th Sarhad J. Agric. Vol.30, No.3, 2014 339 concentrations, respectively. The mortality rate was found acceptable. Toxicity of Tracer was found normal because it did not kill termite workers on spot. They can give time to termites to go back to the colony and allowed feeding their nest mates. Tracer gave required percentage mortality, therefore the Tracer was the best insecticide among all tested insecticides. Our findings are in accordance with those of Sattar (2002) who found that Tracer was found very effective as a slow acting toxicant against H. indicola, C. heimi, M. championis and O. lokamandi. Match (Lufenuron–5% EC) acts as an Insect Growth Regulators (IGRs). Rather than being the typical poison that attacks the insect nervous system they interfere with chitin synthesis and are taken up more by ingestion than by contact. Their greatest values are in the control of caterpillars and larvae. In the present study, the effectiveness of Match as a slow acting toxicant was slightly better by using the highest concentration as compared to the four lower concentrations, which showed the lowest mortality. The average mortality recorded after a period of ten days, i.e. 49.40, 31.06, 26.19, 18.05 and 10.66% from 1st to 5th concentrations, respectively. So the mortality rate was found below from the actual rate. Accordingly, Match proved to be less effective as a slow acting insecticide for controlling H. indicola. These results are in agreement with the findings of Neil et al. (2005), who observed that the effectiveness of Match was not fit as a slow acting toxicant against subterranean termites.

CONCLUSION AND RECOMMENDATIONS Based on the recorded data in the present study the last two concentrations (0.000039% and 0.0000195%) of Regent, the 1st - 3rd concentrations of the Tracer (0.000312, 0.000156 and 0.000078%), and the 1st concentration of the Match (0.00039%) were recommended as slow acting toxicants against H. indicola for keeping them in the bait block station.

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