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Psyche

Volume 2016, Article ID 9417496, 9 pages

http://dx.doi.org/10.1155/2016/9417496

## Functional Responses of* Nephus arcuatus* Kapur (Coleoptera: Coccinellidae), the Most Important Predator of Spherical Mealybug* Nipaecoccus viridis* (Newstead)

^{1}Department of Plant Protection, Faculty of Agriculture, Shahid Chamran University of Ahvaz, Ahvaz, Iran^{2}Department of Plant Protection, College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran

Received 24 September 2015; Revised 27 January 2016; Accepted 3 March 2016

Academic Editor: Juan Corley

Copyright © 2016 Sara Zarghami et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

#### Abstract

*Nephus arcuatus* Kapur is an important predator of* Nipaecoccus viridis* (Newstead), in citrus orchards of southwestern Iran. This study examined the feeding efficiency of all stages of* N. arcuatus* at different densities of* N. viridis* eggs by estimating their functional responses. First and 2nd instar larvae as well as adult males exhibited a type II functional response. Attack rate and handling time were estimated to be 0.2749 h^{−1} and 5.4252 h, respectively, for 1st instars, 0.5142 h^{−1} and 1.1995 h for 2nd instars, and 0.4726 h^{−1} and 0.7765 h for adult males. In contrast, 3rd and 4th instar larvae and adult females of* N. arcuatus* exhibited a type III functional response. Constant and handling time were estimated to be 0.0142 and 0.4064 h for 3rd instars, respectively, 0.00660 and 0.1492 h for 4th instars, and 0.00859 and 0.2850 h for adult females. The functional response of these six developmental stages differed in handling time. Based on maximum predation rate, 4th instar larvae were the most predatory (160.9 eggs/d) followed by adult females (84.2 eggs/d). These findings suggest that* N. arcuatus* is a promising biocontrol agent of* N. viridis* eggs especially for 4th instar larvae and adult females.

#### 1. Introduction

The spherical mealybug,* Nipaecoccus viridis* (Newstead) (Hemiptera: Pseudococcidae), is one of the most important citrus pests in southern and southwestern Iran [1]. This polyphagous pest attacks over 193 plant species throughout tropical and subtropical regions and a large part of the Pacific Basin [2–4]. Chemical control of* N. viridis*, as with other mealybugs, often becomes ineffective due to their cryptic life style in protected locations as well as the presence of a mealy wax that covers its eggs and body [5]. Therefore, biological control using natural enemies has the potential to be an effective alternative method to manage this pest [6–8].

The coccidophagous coccinellid,* Nephus arcuatus* Kapur (Coleoptera: Coccinellidae), is a newly recorded predatory beetle indigenous to the warmer regions of Iran [9]. Until recently, it had only been reported in Yemen and Saudi Arabia [10]. This small coccinellid occurs widely and abundantly in citrus orchards in Dezful, southwestern Iran (personal observation). Recent investigations on the biology and consumption capacity of this predator confirm its potential for the control of* N. viridis* in the citrus orchards [11, 12]. However, more studies are needed to develop this predator within a successful biological management programme.

Prior to using a natural enemy in a biological control programme, it is essential to evaluate its predatory capacity. One of the criteria for determining the efficiency of a predator is the ability of the predator to change its feeding behaviour in response to changes in prey density, that is, its functional response, defined as the number of prey eaten per predator as a function of prey density [13, 14]. Several types of functional response curve have been described, including a linear increase (type I), an increase decelerating to a plateau (type II), or a sigmoidal increase (type III) in which predators cause a constant (I), negative (II), or positive (III) density-dependent mortality of their prey [13, 15, 16]. The functional response curve can be described by evaluating two parameters, the coefficient of attack rate () and the handling time (). The coefficient of attack rate estimates the steepness of the increase in predation with increasing prey density and the handling time helps estimate the satiation threshold [16]. Information on these variables can provide insights into the efficiency of a predator in regulating prey populations, clarifying evolutionary relationships, and predicting the predator’s effectiveness as a biological control agent [16–18].

This study aimed to determine the relative efficiency of different larval instars and of both female and male adults of* N. arcuatus* as biological control agents of* N. viridis*. We achieved this by evaluating the effect of* N. viridis* density on the number of prey consumed by each life stage of* N. arcuatus* to determine the shape of their functional response to prey density, their attack rate coefficients, and handling times.

#### 2. Materials and Methods

##### 2.1. Prey and Predator Cultures

*N. viridis* mealybugs were collected from* Citrus sinensis* L. trees in an orchard in Dezful (48°30′E, 32°20′N), Khuzestan Province, southwestern Iran, in the autumn of 2011. They were then mass-reared on sprouting potato (*Solanum tuberosum* L.) shoots, in rearing boxes (24 × 16 × 10 cm) that were tightly covered by a fine mesh net.* N. arcuatus* adults were collected from the same orchard and reared on sprouted potatoes infested with* N. viridis* for two generations before being used in experiments. The stock colonies of both* N. arcuatus* and* N. viridis* were maintained in an incubator at °C, % RH, and 14L : 10D photoperiod.

##### 2.2. Functional Response Assessments

To obtain a cohort of* N. arcuatus* for experiments, 50 pairs of adult* N. arcuatus* were transferred from the stock culture into a colony of* N. viridis* (mixed developmental stages on 10–12 sprouted potato plants) in a plastic box (20 × 13 × 8 cm) covered with a fine mesh net for ventilation; predator oviposition was allowed to proceed for 12 h after which time the adult predators were removed. Developing predator larvae were observed every 12 h and, over time, developed into cohorts of 1st, 2nd, 3rd, and 4th instar larvae and mated adults males and females (10-day-olds) for use in experiments. Before each developmental stage was evaluated, replicate individuals were kept without food for 12 h in a micro tube (1.5 mL) in order to standardize their hunger level. Thereafter, each predator was introduced into a plastic container (9 × 7 × 3 cm) containing different densities of eggs of* N. viridis* which were the preferred prey for the developmental stage of* N. arcuatus* [19]. Each container had a 20 mm diameter hole in the middle of the lid, which was covered by a piece of fine net to provide ventilation. The densities of* N. viridis* eggs were as follows: 2, 4, 6, 8, 10, 14, and 18 eggs for 1st instar larvae; 2, 4, 8, 16, 20, 30, 40, and 50 eggs for 2nd instar larvae; 2, 4, 8, 16, 20, 40, 60, 80, 100, and 120 eggs for 3rd instar larvae; 2, 4, 8, 16, 32, 60, 100, 140, 180, and 220 eggs for 4th instar larvae; 2, 4, 8, 16, 40, 65, 90, and 115 eggs for adult females; and 2, 4, 8, 16, 20, 35, 50, 60, and 80 eggs for adult males. These densities were selected based on preliminary tests of the consumption capacity of different stages of* N. arcuatus*. After 24 h, predators were removed and the number of eggs consumed was recorded. There were between 10 and 21 replicates for each treatment; greater replication was used for some prey densities to achieve precise information. Experimental conditions were based on optimal temperature for* N. arcuatus* activity: °C, % RH, and 14L : 10D photoperiod [11].

##### 2.3. Statistical Analysis

The functional responses of* N. arcuatus* were analyzed in two steps [20]. In the first step, the type (shape) of functional response was described by determining how well the data fitted to a type I, II, or III functional response, using a polynomial logistic regression of the proportion of prey consumed () as follows:where is the number of prey consumed, is the initial prey density, and the parameters , , , and are the constant, linear, quadratic, and cubic parameters related to the slope of the curve. The above parameters were estimated using the CATMOD procedure in SAS software [20, 21]. The data sets for each developmental stage of* N. arcuatus* were fitted individually to (1) and the types of functional response were determined by examining the signs of and . If was positive and was negative, a type III functional response was evident. However, if was negative the functional response type was a type II [20].

In the second step, a nonlinear least squares regression (PROC NLIN [21]) was used to estimate the functional response parameters ( and either for type II functional response or , , and for type III functional response) using Rogers’s random predator equation which is the most appropriate type II functional response in situations with prey depletion [22]:where is the total time that predator and prey are exposed to each other (24 h); is the attack rate; and is the handling time in hours [20, 23].

For modeling the type III functional response, attack rate () in (2) was substituted in (3) with a function of prey density [16, 24]. In the simplest generalized form, attack rate (3) is a function of the initial number of prey:where , , and are constants that must be estimated. The simplest form arises when is a function of initial density, as in The functional response parameters for 1st instar and 2nd instar larvae and adult males were obtained using (2) (for type II). However, the functional response parameters for 3rd instar and 4th instar larvae and adult females were obtained using (3) and (4) (for type III).

Differences in estimates of attack rates and handling were analyzed using (5) (type II) or (6) (type III) with an indicator variable as follows [20]: where is an indicator variable that takes the value 0 for the first data sets and the value 1 for the second data sets. For a type II response (5), the parameters and estimate the differences between the data sets in the values of the parameters and , respectively. Specifically, the attack rate for one stage is , and that for another stage is . If the parameters and are significantly different from zero, then and , for the two data sets, are different. For the type III response (6), the parameters and estimate the differences between the two data sets being compared with the values of and , respectively. Specifically, the handling time for one stage is , and that for another stage is [20].

The maximum predation rate (), which represents the maximum number of prey that can be consumed by an individual during 24 h, was calculated using the estimated [25].

#### 3. Results

The polynomial logistic regression analysis of the proportion of* N. viridis* consumed by 1st and 2nd instar larvae and by adult male* N. arcuatus* yielded estimated parameters that indicate a type II functional response for these predator stages (Table 1). The linear coefficient was negative for these stages; that is, the proportion of prey consumed declined monotonically with an increase in the initial number of prey offered, which indicates a type II functional response (Figures 1 and 2). Therefore, (2) was used to estimate and . Estimated parameters showed that the 1st instar larva of* N. arcuatus* had the smallest attack rate and handling time compared with 2nd instar larva and adult males (Table 2). The asymptotic 95% confidence interval for included 0 but that for did not, which means that there is a significant difference between and and that these three groups have a different functional response, with a significant difference in handling time, but not in attack rate (Table 3).