Effects of Ethanolic Extract of the Leaves of Pongamia glabra and Gliricidia sepium against Rhipicephalus ( Boophilus ) annulatus

| The effects of ethanolic extracts of the leaves of Pongamia glabra and Gliricidia sepium against Rhipicephalus (Boophilus) annulatus were studied. Different dilutions of the extracts such as 50, 60, 70, 80, 90 and 100 mg/mL were tested using adult immersion test (AIT). The per cent adult mortality, inhibition of fecundity and hatching of laid ova were studied. Both extracts caused very low per cent adult tick mortality and inhibition of fecundity. Fifty per cent blocking of the hatching of laid ova in ticks treated with ethanolic extract of leaves of P. glabra was observed.


IntroductIon
T ick control throughout the world is based mainly on the use of chemical acaricides.Their indiscriminate and repeated use has already resulted in problems related to environmental pollution, milk/meat contamination and the development of resistance in target species along with a subsequent increase in cost (Dipeolu and Ndungu, 1991;Jonsson and Piper, 2007).The use of extracts from various plants is being explored as an alternative to chemical acaricides because they are potentially less toxic to the animals and safer for the environment.

R. (B.
) annulatus, one of the most important bovine one host tick species (Onofree et al., 2001), serves as the vector for the agents of animal as well as some human diseases.R. (B.) annulatus is the major one host tick species of southern India ( Jagannath et al., 1979;Rajamohanan, 1982;Koshy et al., 1982).Present study focuses mainly on the effect of the ethanolic extract of the leaves of P. glabra and G. sepium against adult female R. (B.) annulatus.

plant mateRial
The leaves of P. glabra and G. sepium were collected from Kalpetta, Wayanad district, Kerala.The plants were identified by a botanist and the voucher specimens were deposited in the herbarium of the Department of Botany, Calicut University, Kerala (Accesssion no; P. glabra-CALI 6628, G. sepium-CALI 6643).

pRepaRation of the plant extRaCtS
The leaves were cleaned and dried in shade at room temperature.Dried plant leaves were finely pulverized using a grinder.The powdered plant material (100g) was used for ethanolic extraction in a soxhlet extraction apparatus attached with rotary vacuum evaporator (Butchi, Switzerland).Solvent was completely removed by drying at room temperature.Required quantity of the extracts was weighed and dissolved in 1 per cent Tween-20 or water for making six different dilutions at the rate of 50 mg/mL, 60 mg/mL, 70 mg/mL, 80 mg/mL, 90 mg/ mL and 100 mg/ mL and were used for acaricidal bioassay.

tiCkS
Fully engorged adult female R. (B.) annulatus were collected from infested animals, washed with water and dried using tissue paper.These ticks were used for adult immersion test (AIT).

adult immeRSion teSt (ait)
Various dilutions (50-100 mg/mL) of ethanolic extracts of the leaves of the plants were tested using adult immersion test (Drummond et al., 1973).A total of 336 numbers of ticks were used for the experiment.Four replicates of six ticks were used for each dilution of the extract.Group of six numbers of ticks were weighed prior to the experiment and they were immersed for 2 minutes in the respective dilution (10 mL) in a 50 mL beaker with gentle agitation.Tween 20 (0.1%) was used as control for P. glabra extract while water was used as control for G. sepium.Ticks were recovered from the solution, dried using tissue paper towels and placed in separate plastic specimen tube (25 X 50 mm).The tubes were incubated at 28 0 C and 80 % relative humidity in a BOD incubator.

adult tiCk moRtality
The specimen tubes were observed for the next 15 days for mortality and the per cent adult tick mortality was determined.

StatiStiCal analySiS
Data were expressed as the mean + SEM.Groups were compared using one-way ANOVA for repeated measurements using SPSS software.Duncan's test was used for post-hoc analysis.A value of P<0.05 was considered significant.

rESultS And dIScuSSIon
The results of adult immersion test using the ethanolic extracts of leaves of P. glabra and G. sepium are shown in Table 1 and 2. The efficacy of the extracts against adult female R. (B.) annulatus was assessed by measuring the per cent adult mortality, inhibition of fecundity and hatching rate.
Ethanolic extract of leaves of P. glabra produced adult tick mortality ranging from 4-16 per cent and inhibition of fecundity 4-15 per cent.Adult tick mortality was very low even at higher concentrations (80-100 mg/mL) tested.Ethanolic extract of P. glabra (50-100 mg/mL) considerably blocked the hatchability of the eggs in comparison to the control.Ethanolic extract of G. sepium also produced adult tick mortality in the range of 4-12 per cent.The inhibition of fecundity was dose dependent ranging from 1.31 to 13.61 per cent.The extract did not affect the hatching of eggs laid by treated females.

Advances in Animal and
The toxic phytochemicals that primarily constitute the secondary metabolites affect the nerve axon and synapses, muscles, respiration and behaviour of the insects (Klocke, 1989).In fact, some of these phytochemicals were used to develop commercial insecticides and could serve as models for new insect control agents (Balandrin et al., 1985).
Both extracts tested in the present study were previously reported for their larvicidal effects against mosquitoe vectors (Shanmugasundaram et al., 2008;Sharma et al., 1998).Deltamethrin, the common synthetic pyrethoid compound used in field conditions for cattle tick control, produced a mean adult mortality of 16.662 ±6.803, inhibition of fecundity of 57.3 per cent and hatching of 0-10 per cent (Sunil et al., 2013;Divya et al., 2014) at 30 ppm concentration.
Similarly, cypermethrin at 200 ppm concentration also caused mortality of 45.83 per cent and inhibition of fecundity of 86.99 per cent, while the drug did not affect hatching of ova laid by treated ticks (Ravindran et al., 2014).
In the present study, the crude ethanolic extracts of the leaves of the P. glabra and G. sepium did not cause considerable adult mortality or inhibition of egg laying capacity of the treated ticks.The extract significantly inhibited the hatching of the laid ova by treated ticks.Our observation

table 1 :
Effect of ethanolic extract of leaves of P. glabra against R. (B.) annulatus

table 2 :
Effect of ethanolic extract of leaves of G. sepium against R. (B.) annulatus n: 4; Values are Mean +SEM; a, b, c (P<0.05): Significant difference when compared with the control