Functional response of Coccinella septempunctata L . ( Coleoptera : Coccinellidae ) on Ommatissus lybicus ( Homoptera : Tropiduchidae )

Dubas bug, Ommatissus lybicus (Homoptera: Tropiduchidae), is a major insect pest of date palm. Adult and nymph stages suck sap from all green parts and cause direct and indirect damage. Large number of predators feed on it; however, no information is available on their consumption rate and functional response using this prey. We aim to find out consumption rate of Coccinella septempunctata with an objective how adult female and fourth nymphal instars of C. septempunctata responds to 1 and 3 instar nymphs of O. lybicus at various density levels under controlled condition. Both stages of C. septempunctata exhibited a type II functional response. Handling times and attack rates were significantly different between adult female and larval stage indicating that predator responds differentially to prey stage offered at same density. Estimated handling time of C. septempunctata adult female and fourth instar (in bracket) at 24 hours were 0.563(0.692) and 1.008(1.492) and estimated attack rate were 0.020 (0.014) and 0.003(0.002) on first and third instar nymphs of prey, respectively, which conclude that adult is more voracious stage than larva. However, further field studies on predator and prey stage specific in the presence of honeydew droplets on leaflets are still needed to quantify biocontrol efficiency of predator.


Frond and cage size selection
Number of fully expanded fronds of Kehraba cultivar with ten (10) middle leaflets; five on each side were selected.Both cut ends (sides) of frond were covered with wet cotton to reduce water losses and keep the turgidity of leaflets during the course of experiment.A single frond was placed in each cages (20x17x3 inch) with a wooden frame and covered by muslin cloth in order to maintain the prey and predator similar to natural/field conditions.

Prey stage selection and culture
Two developmental stages (1 st and 3 rd instar nymphs) of dubas bug (prey) were chosen.In order to test the predation rate and determine the functional response of both stages of predator on each developmental stage of prey at a time, a number of 1 st instar nymphs of dubas bug were collected with aspirator and maintained as stock population.When more than 90% of the stock population of prey reached to 3 rd instar, required numbers of 1 st instar nymphs were collected from field and offered to the predator.Similarly, stock population of C. septempunctata was maintained on Medicago sativa and fed with aphids.Estimating consumption rate and functional response Consumption rate and functional response of fourth instar and adult female of C. septempunctata were determined by offering the 1 st and 3 rd instar nymphs of dubas bug at different density levels (1, 5, 10, 20, 40, and 60 bugs per leaflet or a total of 10, 50, 100, 200, 400 and 600 bugs per cage).Each life stage of predator and density level of prey was released randomly into the single cage to each predator stage.Similarly, control cages received corresponding prey density level and developmental stages but no predator.Both stages of predators were starved for 24 hours before introducing into the cages.Exposure time of prey to predator was 24 hours and without replacement of prey.Following 24 hours, mean number of killed/consumed prey were counted in both treated and control cages.The experiment was conducted controlled conditions (temperature 30±2 °C, R.H. 20±2 % and photoperiod 16L:8D) replicated five times.

Data analysis
The logistic regression model [19] was used to determine the shape of the functional response prior to fitting the data for functional response analysis.This model helps distinguishing between Type II and Type III [17].The relationship between the proportion of prey consumed (Ne / N0) and number of prey present (N0) was determined by fitting the data to a polynomial function (equation 1) using statistical software SAS, Version 9.0 (Procedure CATMOD) on personal computer.
Where Po, P1, P2, and P3 were the intercept, linear, quadratic and cubic coefficients, respectively, which were estimated using maximum likelihood method.If P1 > 0 and P2 < 0, the proportion of prey consumed is positively density dependent, thus describing a type III functional response.If P1 < 0, the proportion of prey consumed declines with initial prey density suggesting a type II functional response [19].Later on, the functional response parameters were estimated using Roger's random predator equation (Rogers 1972).For Type II response the equation will be followed: Where, Ne is number of prey consumed and N0 is initial prey density, a is the attack rate, T is the total time of exposure, and Th is the handling time.Random predator equation overcomes the problem of prey depletion.Data fitting was performed using nonlinear least square method on SAS Version 9.0 (Procedure NLIN) [19].For Type III response equation will be followed: where b, c and d are constants.The data on prey consumption by fourth instar and adult female C. septempunctata at different prey stage level were subjected to one-way ANOVA and the comparison of means was done using Tukey's HSD test using statistical software, SAS.The number of first and third instar nymphs of dubas bug at different density levels consumed by fourth instar and adult female of C. septempunctata in 24 hours were analyzed using two-way ANOVA by using "prey stage" and "prey density" level as independent factor and "prey consumed" as dependent factor using statistical software, SAS.Comparative consumption rate of fourth instar and adult female C. septempunctata Significant difference was observed in the predation rate of adult female and fourth instar larva of C. septempunctata feeding on the same density and prey stage with same environmental conditions.Irrespective of prey density, mean predation per 24 hours of adult and grub stages of C. septempunctata was recorded as 13.66 and 12.48 prey, respectively (Fig. 3).

Functional response
Parameter estimated for logistic regression of proportion of prey killed (Ne/No) against the number of prey offered (No) for the adult female and 4 th instar grub of C. septempunctata feeding on first and 3 rd instar nymphs of dubas bug displayed type II functional response, i.e. significant linear parameter P1 < 0 (table 3 and fig. 1 and 2). Figure 1 and 2 illustrated that functional response curves and predation percentage of both developmental stages of C. septempunctata was significantly high feeding on 1 st than 3 rd instar nymphs of dubas bug.Similarly, irrespective of prey stage the overall functional response curve of adult was higher than the grub, whereas at higher prey density the prey consumed by both developmental stages was at par (fig.3).Two-way ANOVA revealed a significant main effect of "prey stage" (F= 314.79;P<0.0001; d.f.= 1) and "prey density" (F= 278.93;P<0.0001; d.f.= 5) when adult female C. septempunctata was searching.The interaction between "prey stage" and "prey density" was also found to be significant (F= 20.87; P<0.001; d.f.= 5).Similarly, main effect of "prey stage" (F= 584.64;P<0.0001; d.f.= 1) and "prey density" (F= 321.23;P<0.0001; d.f.= 5) was also significant when fourth instar C. septempunctata was searching.The interaction between "prey stage" and "prey density" was also found to be significant (F= 37.71; P<0.001; d.f.= 5).species, size, development stage, degree of predator hunger and most importantly some environmental variables.Search rate (a) and handling time (Th) of both stages of C. septempunctata was significantly high preying on 1 st instar nymphs of dubas bug compared to 3rd instar nymphs; indicated comparatively more mobility of 3 rd instar than 1 st instar dubas bug.Other possibly reasons may be due to variation in prey distribution [28], effect of temperature on predation and search rate of predator [37, 38].While higher handling time of both stages on 3 rd instar nymphs showed that both stages of predator need more time to grasp 3 rd instar than 1 st instar.In comparison, adult had significantly shorter handling time and higher search rate compared to grub, they take less time to process the food and take comparatively more time to locate the prey (may be due to mobility), which showed that C. septempunctata adult is more efficient in terms of food processing.Based on present result of consumption rate, functional response, handling time and attack rate between two stages of predator that varied significantly we can conclude that though both development stages of C. septempunctata illustrated type II functional response but adult is the most efficient than grub.Functional response of predators can be affected by different biotic and a-biotic factors.However, field studies on predator and prey stage, sex specific, temperature and host plant in the presence of honeydew droplets on leaflets are still needed to quantify biocontrol efficiency of predator.

Figure 1 .
Figure 1.Functional response of adult female and fourth instar C. septempunctata (obtained from the predicted values of prey consumed by PROC-NLIN procedure of SAS), preying upon 1 st and 3 rd instar nymphs of dubas bug offered at different density levels (average of four seasons, 2009-10)

Consumption Rate (average of four seasons) C. septempunctata (adult female) vs. Dubas bug first and third instar nymph
Consumption rate of adult female C. septempunctata fed on first (F=162.98;P<0.0001; d.f.= 5, 29) and third (F=121.19;P<0.0001; d.f.= 5, 29; Table-1) instars of dubas bug increased significantly with increase in prey density levels.Comparison of means revealed that prey consumption was significant at all the density levels except at prey densities, 400 and 600 of first instar nymph when adult female was used as predator (Tukey's range = 4.31).C

. septempunctata (grub) vs. Dubas bug first and third instar nymph
Almost similar trend was observed when fourth instar C. septempunctata was used as predator at different densities of first and third instars of dubas bug (table2 and figure.1).Prey consumption by fourth instar of C. septempunctata increased significantly with increase in densities of first (F=205.82;P<0.0001; d.f.= 5, 29) and third (F=121.93;P<0.0001; d.f.= 5, 29) instar nymphs of dubas bug.Mean values also varied significantly at all the density levels except at prey densities, 400 and 600 of first instar nymph when fourth instar C. septempunctata was used as predator (Tukey's range = 4.31).

Table 1 . Mean number (±SE) of first and third instar nymphs of O. lybicus, consumed (in 24 hours) by adult female C. septempunctata at different prey densities (average of four seasons: spring and summer, 2009-10)
*Means followed by different letters in the same column are significantly different at P<0.001; Tukey's Range = 4.31)

Table 2 . Mean number (±SE) of first and third instar nymphs of O. lybicus, consumed (in 24 hours) by fourth instar C. septempunctata at different prey densities (average of four seasons: spring and summer, 2009-10) Treatment Dubas bug development stage Density/cage First instar Third Instar
*Means followed by different letters in the same column are significantly different at P<0.001; d.f.= 5, 29; Tukey's Range = 4.31)

Comparative functional response of C. septempunctata adult and grub at different densities of dubas bug (irrespective of life stages) Handling time and search rate
Prey stage significantly affected the handling time and it increased with increasing prey size.Adult female C.