MATERIALS AND METHODS

Collection of plant oils: Clove (Syzygium aromaticum Linn.), citronella (Cymbopogon nardus var. Linn.), citriodora (Corymbia citriodora var. citriodora), Gaultheria (Gaultheria procumbens Linn) and lemongras (Cymbopogon citratus) were purchased from Tamil Nadu Government Co-operative Super Market, Cherring cross, Udhagamandalam, The Nilgiris, Tamil Nadu, India and collected oils were used for bioassay against larvae of T. castaneum.

Culture of test organism: Tribolium castaneum larvae was collected from infested grains purchased from local market in Musiri, Tamil Nadu, India and brought to the laboratory. The culture was established using wheat flour in a plastic container of 25×10 cm and maintained at room temperature 30±2°C and relative humidity of 70-75%. Sieving the culture separated the 4th instar larva and the larvae were used for subsequent experiment. The culture was continuously maintained in the containers throughout the study period.

Repellent activity: Repellent activity contained in the five essential oils were studied according to method adopted by Talukder and Howse (1994). Petridishes (9 cm dia) were used for the repellency test. Test solutions were prepared by dissolving different concentrations (5, 10, 15 and 20 μL) of each oil in 1 mL distilled water. Whatman No. 1 filter paper (8 cm dia) was cut into two and each solution was applied to half of a filter paper as uniform as possible by using micro pipette. The other half of the filter paper was treated with distilled water alone. The oil treated and distilled water untreated halves were dried to evaporate the solvent completely. Treated and untreated halves were attached with staple pins and placed in the glass petri dish. About 10 No. of 4th instar larvae of T. castaneum were released at the centre of the filter paper disc and then sealed tightly. Five replicates were set for each concentration. Observation of the No. of larva present on both the treated and untreated halves were recorded after 30 min for 2 h of experiment setting (30, 60, 90 and 120 min). The data were expressed as Percentage Repulsion (PR) by using the following equation (Talukder and Howse, 1994):

where, PR is percentage repellency, NC is No. of larvae on control portion, NT is No. of larvae on treated portion.

The average values was then classified according to repellency classes from 0-V, where 0 = <0.1, Class I = 0.1-20, Class II = 20.1-40, Class III = 40.1-60, Class IV = 60.1-80 and Class V = 80.1-100%, respectively (McGovern et al., 1977).

Larvicidal activity: For evaluation of larvicidal activity in newly moulted 4th instar larvae of T. castaneum were exposed to various quantities of essential oils. To study the larvicidal activity of each oils are 3 g of half broken rice was coated with different concentrations (5, 10, 15 and 20 μL) in 1 mL of distilled water placed in (9 cm dia) glass petri dish and distilled water was used as control. The oil treated and distilled water untreated was dried to evaporate the solvent completely. Ten larvae taken from laboratory culture were placed in each petri dish. Five replicates were set for each concentration and control. Mortality was recorded every 24 h for 4 days and the percentage of mortality was calculated by Abbott (1925):

Statistical analysis: Data analysis was carried out using Microsoft Excel 2007. One-way ANOVA was performed for all the experimental data from that least significant difference was calculated and the significant differences were marked with different alphabet.

RESULTS

Repellent activity of five essential oils was studied at four different concentrations (5, 10, 15 and 20 μL mL^–1) against T. castaneum larvae. Table 1 is showed remarkable repellent effects. Repellency was seen in the number of larva found in the treated oils were significantly smaller than the number in the control.

Table 1:	Repellent activity of five essential plant oils against T. castaneum larvae
ND: Not Detected, 0 = <0.1, Class I: 0.1-2, Class II: 20.1-40, Class III: 40.1-60, Class IV: 60.1-80 and Class V: 80.1-100%

Table 2:	Larvicidal activity of five essential plant oils against T. castaneum larvae

Higher concentration (20 μL mL^–1) of all oils showed higher repellency in T. castaneum larvae. Data pertaining to the above experiment clearly revealed that maximum repellent activity was recorded in citriodora oil 95.24% on T. castaneum larvae at 20 μL mL^–1 concentration compared to control. Following descending order shows the repellency of other four oils in T. castaneum larvae at concentration (20 μL mL^–1) Gaultheria oil (80.2%)> lemongrass oil (71%)> citronella oil (63%)> clove oil (61%), respectively.

Larvicidal activity of five essential oils were tested against T. castaneum larvae at four different concentrations. Data pertaining to the larvicidal activity of the selected plant oils was presented in Table 2. The results indicating that variation among the plant oils tested against the selected insect pest. Toxicity increase with increasing concentration, exposure period and insect species, indicated that larvae were significantly susceptible to the five essential oils after 24-96 h of treatment. Larvicidal activity caused maximum toxicity was recorded in citriodora oil 81.86% at the rate of 20 μL mL^–1 concentration. Gaultheria oil showed 69.91% mortality at 96 h after treatment. Whereas lemongrass oil, citronella oil and clove oils showed moderate activity compared with citriodora oil. One-way analysis of variance (ANOVA) followed by Least Significant Difference (LSD) test showed statistical significance (p<0.05) compared to control.

DISCUSSION

Plant products having considerable potential as insecticidal compounds are gaining tremendous importance in recent years. Approximately two-thirds to three-quarters of the world’s population rely on medicinal flora as their main source of medicines (Anonymous, 2000). Many studies were reported on the insecticidal activity of essential oils against rust-red flour beetle and dried bean beetle (Shaaya et al., 1991; Pemonge et al., 1997; Papachristos and Stamopoulos, 2004; Mondal and Khalequzzaman, 2006; Chaubey, 2007; Islam et al., 2009).

In the present investigation larvae of T. castaneum were found to be significantly repellant at concentration of 20 μL mL^–1 of citriodora oil in compared with other four oils and control. These results are consistent with the earlier reports repellent activity of plant derived essential oils, such as bael (Aegle marmelos), dhaniya (Corriandrum sativum), laung (Schyzygium aromaticum) and orange (citrus reticulata) against Sitophilous oryzae and T. castaneum. Highest repellant activity was observed in S. aromaticum against S. oryzae (90%) and T. castaneum (90%) (Mishra and Tripathi, 2011). Mahmoodavand and Shakarami (2014) noticed repellency effect of essential oils and powders of Mentha longifolia, Thymus daenensis, Achillea wilhelmisii and Artemisa haussknechtii against adults of T. castaneum and T. confusum. The essential oil of M. longifolia exhibited significantly stronger repellency effects as at 0.9 μL oil concentration caused 86.7 and 80% repellency of T. castaneum and T. confusum, respectively. Caballero-Gallerdo et al. (2012) indicated that adult beetles of T. castaneum were repelled significantly by essential oils of Cymbopogon martini, C. flexuosus and Lippia origanoides. Citronella oil also used to mosquito repellent qualities has been verified by research (Kim et al., 2005). Moore et al. (2007) also reported that use of essential oil of lemongrass as a repellent in Anopheles darlingi and Anopheles arabiensis.

In the present study citriodora oil exhibited significant larvicidal activity at higher concentration. It is possible that the insecticidal property present in the selected plant compound may arrest the various metabolic activities of the larvae during the development and ultimately the larvae failed to moult and finally died. Mortality increased with increasing of both concentration and exposure period. This results coincide with the earlier findings of Khani and Asghari (2012) who noticed that insecticidal activity of essiential oils of Mentha longifolia, Pulicaria gnaphalodes and Achillea wilhelmsii against T. castaneum and Callsobruchus maculatus. These three oils showed strong insecticidal activity against C. maculatus but M. longifolia and A. wilhelmsii oils showed strong larvicidal activity against T. castaneum. Popovic et al. (2013) reported that essential oils from three plants against T. castaneum. Powerful larvicide and repellent effect was observed that essential oils of Calamintha glandulosa with concentration of 1.14% showed higher mortality rate after 24 h (56.67%), respectively. Jeyasankar (2012) already reported that antifeedant, insecticidal and growth inhibitory activities of selected plant oils on black cutworm, Agrotis ipsilon. Maximum percentage of insecticidal activity (86.92%) and deformities in treated larvae was recorded in gaultheria oil. Clove oil is also used to control insects, fungus, mildews in stored grains (Han et al., 2006; Shang and Cai, 2007). Some authors reported that citronella oil against stored grain pests Some plant extracts, essential oils or their constituents described in this review have demonstrated high efficacy against coleopteran stored products insect pests responsible for post-harvest damage.

CONCLUSION

Citriodora oil showed greater performance of larvicidal and repellent activities against T. castaneum. Hence, it may be suggested that the citriodora oil, can be used for controlling the insect pest, T. castaneum, which will replace the chemical pesticides.