Systematic & Applied Acarology Special Publications (2000) 4,

Systematic & Applied Acarology Special Publications (2000) 4, 109-124 109 Potential of Amblyseius cucumeris (Acari: Phytoseiidae) as a biocontrol agent against Schizotetranychus nanjingensis (Acari: Tetranychidae) in Fujian, China YANXUAN ZHANG 1, ZHI-QIANG ZHANG 2, JIANZHEN LIN 1 & JIE JI 1 1 Institute of Plant Protection, Fujian Academy of Agricultural Sciences, Fuzhou 350013, China; E-mail: zyxlj@pub3.fz.fj.cn 2 Landcare Research, Private Bag 92170, Auckland, New Zealand Abstract The predatory mite Amblyseius cucumeris (Oudemans) was studied in the laboratory to evaluate its potential as a biocontrol agent against the spider mite Schizotetranychus nanjingensis Ma & Yuan, a pest of the moso bamboo in Fujian, China. When fed S. nanjingensis females and eggs, the life cycle of A. cucumeris (developmental time from egg to egg 7.7 days for the first generation and 7.8 days for the second generation) was as long as its life cycle on its normal diet in the laboratory, Tyrophagus putrescentiae (Schrank) (7.8 days) at 27-28 o C. Adult females of A. cucumeris started to lay eggs at the age of 3 days with a daily rate of 1-4 eggs (average of 2.2) over a period of 7-18 days and a total fecundity of 35.8 (14-47) eggs. The number of prey consumed by predators increased with prey density and the number of eggs produced was directly correlated with the number of prey consumed. Female predators consumed twice as many female spider mites as did male and deutonymph predators (6 versus 3 per day at 9 prey per leaf). A. cucumeris females were unable to invade intact webnests of S. nanjingensis but were able to invade and liked to stay and lay eggs in broken nests with existing openings or holes. Key words: Acari, spider mites, predatory mites, biocontrol, bamboo

110 Systematic & Applied Acarology Special Publications (2000) 4 Introduction The spider mite Schizotetranychus nanjingensis Ma & Yuan is one of the phytophagous mites often causing economic injury to the moso bamboo (Phyllostachys pubescens) in Fujian province, China (Zhang et al. 1998ab, 1999b). This species constructs nests with a very dense web roofing on the underside of bamboo leaves. The webnest functions as a shelter within which all mites live and feed on leaf tissue. The upperside of an infested leaf is full of whitish stippling in the early stages of mite infestation. As mite feeding continues, leaves become yellowish and brown. Looking from a distance, a seriously infested bamboo forest has a "burnt" appearance, with leaves falling and photosynthesis of the plant severely reduced (Yu & Shi 1991; Zhang et al. 1998a). In bamboo forests, Typhlodromus bambusae Ehara is a natural enemy of S. nanjingensis and other phytophagous mites (Zhang et al. 1999c). However, its density is low in the growing season and can not provide natural control of phytophagous mites in many forests. Another predator, A. longispinosus Evans, is also known to occur in very low densities on bamboo in Fujian. It was evaluated as a possible biocontrol agent (Zhang et al. 1998c) and mass release was also made (Zhang et al. 2000) in bamboo forests. However, this species is relatively expensive to rear. We therefore feel it is necessary to search for a predator which is easy and cheap to produce. The predatory mite Amblyseius cucumeris (Oudemans), sometimes cited in literature as Neoseiulus cucumeris, is such a predator. It is a biocontrol agent widely used for the control of various species of thrips on cucumber and pepper in greenhouses through preventive, mass releases (Gillespie 1989; Ramakers et al. 1989; Bennison & Jacobson 1991; Higgins 1992; Workman et al. 1994; Jacobson 1995; Cheyrias 1998; Larsen et al. 1998; Wada 1999). It has also been used to control thrips on ornamentals in greenhouses with varying degrees of success

ZHANG et al.: Amblyseius cucumeris against Schizotetranychus 111 (Wardlow et al. 1991; Agostinelli et al. 1993; De Courcy Willians 1993; Hessein & Parrella 1993; Stanton 1994). Although widely used in greenhouses, this predator is also found naturally in the field (Workman et al. 1994; Iraola et al. 1997) and was recently used for thrips control in field crops (Dissevelt et al. 1995; Schade & Sengonca 1995; Bennison et al. 1996). Interestingly, this phytoseiid has rarely been used against phytophagous mites. One reported evaluation of it against the eriophyid mite Aculops lycopersici (Massee) showed that it failed to reproduce on this mite (Brodeur et al. 1997). A. cucumeris was effective against the broad mite Polyphagotarsonemus latus (Banks) on greenhouse plants in China (L.R. Liang, personal communication). But so far A. cucumeris has not been used for spider mite control. This study examines its potential as a predator against S. nanjingensis. Materials and methods Mites and plants The predatory mite A. cucumeris was introduced from Biological Crop Protection Limited in England and reared using Tyrophagus putrescentiae (Schrank) in wheat bran in the laboratory. The first group of S. nanjingensis used in this study originated from the moso bamboo in Gongchuan Village, Yongan county, Fujian province, China. The second group of leaves of moso bamboo with or without mites were collected from bamboo forests in Nanping county. Leaves not immediately used were stored in platic bags at 5 o C. General description of the experimental setup Petri dishes (diameter 6 or 12 cm) were used as platforms for rearing and observing mites (see Zhang et al. 1998c for details). Bamboo leaves were placed on a piece of filter paper which rested on a piece of foam plastic soaked with water in the Petri dish. Leaves were replaced every five days by placing a fresh leave near to the old leave. When the mite moved to the new leaf, the old leaf was removed and discarded.

112 Systematic & Applied Acarology Special Publications (2000) 4 Development An egg of the predatory mite was transferred to a clean bamboo leaf. The development of the egg was followed by observations at 12- hour intervals until it reached adulthood. The predatory mites were fed spider mites by adding 4 adult females per day in the rearing unit. The first generation from eggs produced by adult predators reared on S. nanjingensis was fed S. nanjingensis again for a second generation and the duration of each life stage was measured as did for the first generation. In another set of experiments designed for comparison, predatory mites were also fed an abundance of T. putrescentiae, which were added to bamboo leaves with some bran. There were 12-18 replicates per treatment. The experiment was conducted at 27-28 o C. Reproduction A newly mated female predatory mite was transferred to a green bamboo (Dendrocalamus latiflorus) leaf portion (2 x 2 cm) and was fed an abundance of S. nanjingensis females and eggs that the spider mite females laid. Oviposition by the female predator was checked daily and eggs laid each day were counted and removed. Observations continued until the predator stopped oviposition, or died. There were 6 replicates. The experiments were conducted in a growth chamber at 28-30 o C. Predation and reproduction in relation to prey density The experiment is the same as the above one except that (1) the experiments were conducted at the following different prey densities: 1, 3, 5, 7 and 9 prey females per leaf, 2) the experiment was continued for 2 days, and 3) the number of prey consumed per day were also recorded.

ZHANG et al.: Amblyseius cucumeris against Schizotetranychus 113 Functional responses of different predator stages The experimental arena was the same as above. Each predator was starved for 12 hours before test and fed 1, 3, 5, 7 or 9 S. nanjingensis females. The numbers of prey consumed were counted at the end of 24 hours. Prey numbers were then made up to the original density and the number of prey consumed counted again after another 24 hours. There were six replicates per density. The experiment were repeated for female predators, male predators and deutonymph predators. Predator interaction with webnests in bamboo leaves Ten leaves of the moso bamboo each with 1-2 intact webnests were each placed on a piece of wet filter paper in a petri dish. Two female predatory mites were then introduced onto each leaf. Observations were then made at 6, 24 and 72 hours after the start of the experiment. The same procedure was applied to another group of ten leaves each with 1-2 broken webnests (naturally broken, with at least one hole on the web). Observations were made in a room at 25-27 o C. Data analyses ANOVAs and regressions were performed using SYSTAT 7.0 (SPSS Inc. 3/1997). Parameters of the functional responses were estimated using the Woolf transformation of the disc equation advocated by Fan and Petitt (1994). Results Development Amblyseius cucumeris eggs, laid by females which were reared on T. putrescentiae, developed normally on S. nanjingensis; immature survival rate was 100%. Predator eggs laid by the first generation of females reared on S. nanjingensis also developed normally; immature survival rate was 94%. The developmental times of each stage were

114 Systematic & Applied Acarology Special Publications (2000) 4 similar when predators were reared on T. putrescentiae and S. nanjingensis (Table 1); the egg stage and pre-oviposition period (each about 2.5 days) were much longer than the larval stage and two nymphal stages, each of which lasted about 1 day at 27-28 o C. TABLE 1. Development of Amblyseius cucumeris on Schizotetranychus nanjingensis for two generations and on its culture diet of Tyrophagus putrescentiae (27-28 o C). Data presented are means±standard errors for n=12-18. Prey types Egg Larva Protonymph Deutonymph Pre-oviposition T. putrescentiae 2.56±0.62 0.71±0.27 1.14±0.24 1.00±0.41 2.43±0.19 S. nanjingensis 1 st generation 2.41 ± 0.12 0.83±0.07 0.96±0.04 1.17 ± 0.07 2.50±0.11 2 nd generation 2.41±0.05 0.84±0.24 0.88±0.06 1.07±0.05 2.60±0.39 Reproduction Amblyseius cucumeris adult females started to lay eggs at the age of 3 days when fed S. nanjingensis (Table 1, Fig. 1). The rate of oviposition increased rapidly during the first 3 days of the oviposition period and then fluctuated between 1-4 eggs (Fig. 1), with an average of 2.2 eggs per day (Table 2). Near the end of the oviposition period (7-18 days), oviposition rate rapidly decreased (Fig. 1). Total fecundity averaged 35.8 (14-47) eggs per female (Table 2). Eggs laid per day Age of adults (day) FIGURE 1. Daily rate of oviposition by Amblyseius cucumeris females feeding on an abundance of Schizotetranychus nanjingensis females at 25-27 o C.

ZHANG et al.: Amblyseius cucumeris against Schizotetranychus 115 TABLE 2. Reproduction of Amblyseius cucumeris fed on Schizotetranychus nanjingensis (25-27 o C). Reproductive traits Mean±SE No. of predators observed Fecundity 35.83±4.59 6 Oviposition period (day) 16.17±1.83 6 Reproductive rate (per day) 2.20±0.09 6 Predation and reproduction in relation to prey density The rate of prey consumption by A. cucumeris females increased significantly with prey density (F 4,25 =75.62; P<0.001; Fig. 2A). The number of eggs produced by these females also increased with prey density (F 4,25 =17.65; P<0.001; Fig. 2B). No. prey consumed (/day) A No. eggs laid (/day) B Prey density Prey density FIGURE 2. (A) Rate of predation and (B) rate of oviposition (per day) of Amblyseius cucumeris females in relation to the density of Schizotetranychus nanjingensis females at 28-30 o C. There was a positive linear relationship between the number of prey consumed (x) and the number of eggs produced (y): y = 0.547 + 0.427 x, r=0.841 (Fig. 3).

116 Systematic & Applied Acarology Special Publications (2000) 4 No. eggs laid (/day) No. prey consumed (/day) FIGURE 3. Relationship between rate of oviposition (per day) and rate of predation (per day) of Amblyseius cucumeris females feeding on Schizotetranychus nanjingensis females at 28-30 o C. Functional responses of different predator stages The number of prey consumed by all three life stages of A. cucumeris increased with prey density, but the magnitude of responses varied among them (F 2,84 =29.86, P<0.001; Fig. 4). Adult females on average consumed about twice as many prey as adult males and deutonymphs (Fig. 4). This was due to the short handling time of adult females compared to adult males and deutonymphs (Table 4). Not surprisingly, successful rate of attack decreased in the order of the size of predatory mites: adult female > adult male > deutonymphs. TABLE 3. Estimated parameters of functional responses of Amblyseius cucumeris to Schizotetranychus nanjingensis (25-27 o C). Life stage of predators Handling time Successful attack rate Adult female 0.062±0.018 1.159 Adult male 0.305±0.040 0.989 Deutonymph 0.207±0.079 0.585

ZHANG et al.: Amblyseius cucumeris against Schizotetranychus 117 A No. prey consumed per day B C Prey density FIGURE 4. Functional responses of adult females (A), adult males (B) and deutonymphs (C) of Amblyseius cucumeris to different densities of Schizotetranychus nanjingensis females at 25-27 o C. Predator interaction with webnests on bamboo leaves None of the 20 predatory mites tested were able to invade the intact webnests of S. nanjingensis in three days; 20% of them escaped from the leaf in 6 hours, 50% escaped in 24 hours and 95% escaped in 72 hours. A few eggs were seen to have been laid on the surface of web by a couple of predators. In contrast, 75% of the predators invaded the broken webnests in six hours and they laid 0.47 eggs per female in the webnest. At 24

118 Systematic & Applied Acarology Special Publications (2000) 4 hours after the start of the experiment, 80% of the predators were in the broken nests and 0.50 eggs per female were found in the nests. By 72 hours, prey in the webnests were all consumed and only 40% of the predators remained in the nests while the others had disappeared; predatory mite nymphs were seen in invaded webnests or on the surfaces of 40% of the leaves. Discussion Although A. cucumeris has never been associated with S. nanjingensis before, we showed in this study that it readily attacked S. nanjingensis, and could develop on this spider mite normally at least as fast as it could on Tyrophagus on which it is normally reared in the laboratory. The rate of reproduction of A. cucumeris on S. nanjingensis (2.2 per day) is higher than those on other prey (1. 4 in Dosse 1955; 1.9 in Castagnoli 1989). These results indicate that S. nanjingensis is quite a good prey for A. cucumeris in terms of nutrition. Our experiments on functional responses also showed that predation rates of A. cucumeris rapidly increased with the density of the prey S. nanjingensis and the number of eggs produced by predators (y) were positively related to the number of prey taken (x), with a y/x slope of 0.427. This is higher than that of A. longispinosus feeding on S. nanjingensis (0.335; Zhang et al. 1999a). This is very interesting as it shows that A. cucumeris is more efficient than A. longispinosus in converting food (S. nanjingensis) to eggs. An important concern of using A. cucumeris against S. nanjingensis is whether it can invade the tightly weaved webnests which are built by S. nanjingensis females to protect themselves and their young against predators and extreme environmental conditions (Zhang et al. 1999b). Our observations showed that A. cucumeris females were not able to invade the intact webnests of S. nanjingensis; however, they liked webnests which had an opening or hole and preferred to stay and lay eggs inside these webnests, using them as their own nests. We

ZHANG et al.: Amblyseius cucumeris against Schizotetranychus 119 observed that predator eggs developed inside these nests. It is important to note that infested bamboo leaves in the field usually have a high proportion of webnests damaged in some way; the damage is caused either by other insects or by leaves rubbing against each other in the wind. Therefore, A. cucumeris can attack a large proportion of S. nanjingensis in these broken nests and can also attack spider mites that are expanding their nests or in dispersal. Another concern is whether A. cucumeris can adapt to foraging on bamboo plants. We released these mites onto branches of bamboo with many leaves infested with webnests and found some of the predatory mites located the webnests and invaded broken ones. Mass-reared predators were also released onto bamboo plants by fixing small boxes of A. cucumeris (with some Tyrophagus and bran) on bamboo culm. The experiments are on-going and results will be reported in the future. It is interesting to note that Fujian has a mild climate and is quite humid. Bamboo forests are relatively stable environments with a relatively high humidity. This is in a way similar to the greenhouse environment, where A. cucumeris does well and is widely and successfully used for biocontrol of thrips on a variety of crops. Acknowledgments This project is a Key Project of the Nine-Five Period sponsored by Fijian Science Council. Zhi-Qiang Zhang's visits to Fujian in July 1997, September-October 1998 and October 1999 were funded by Fujian "Yinzhiban", a provincial office for promoting exchange and collaboration between scientists in China and elsewhere. This paper was prepared and finalized when the junior author was funded by the Foundation for Research, Science, and Technology, New Zealand, under contract number C09617. We thank Dr Anne Baker (The Natural History Museum, London) for a review of the manuscript.

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