- Email: [email protected]
Effects of combining releases of non-viable host eggs with insecticide application on Riptortus pedestris population and its egg parasitoids
Journal of Asia-Pacific Entomology 15 (2012) 299–305
Contents lists available at SciVerse ScienceDirect
Journal of Asia-Pacific Entomology journal homepage: www.elsevier.com/locate/jape
Effects of combining releases of non-viable host eggs with insecticide application on Riptortus pedestris population and its egg parasitoids Bishwo P. Mainali a, Sangwon Kim b, Un Taek Lim a,⁎ a b
School of Bioresource Sciences, Andong National University, Andong 760–749, Republic of Korea Dongbu Ceres Co. Ltd., 135 Dongsan, Nonsan, Republic of Korea
a r t i c l e
i n f o
Article history: Received 8 December 2011 Revised 24 March 2012 Accepted 31 March 2012 Keywords: Ooencyrtus nezarae Gryon japonicum Cold storage Parasitism Thiamethoxam Alydid
a b s t r a c t Our previous study demonstrated that the release of refrigerated non-viable eggs of Riptortus pedestris (Fabricius) (Hemiptera: Alydidae) enhanced parasitism rates in soybean fields but did not result in the reduction of R. pedestris populations. This study was further conducted using an open-cage exclusion design in a soybean field in order to evaluate the compatibility of combining releases of non-viable host eggs with a single pre-harvest application of insecticide for the control of R. pedestris. Refrigerated eggs of R. pedestris were released twice in treatment plots, and fresh (b 1 day old) eggs of R. pedestris were deployed in all experimental arenas, every 6 days, for host resource and measurement of field parasitism. The releases of host eggs did not reduce the number of R. pedestris in any life stage except the adult stage on two sampling dates. However, parasitism by Gryon japonicum (Ashmead) (Hymenoptera: Scelionidae) was higher in treated plots (9–25%) than in the control plots (1–9%). Statistical significant reduction was not found in the pest population, but parasitism rates significantly increased. Pesticide application did not reduce the bug population but did affect the parasitoids population. Pest management tactics, using both artificially deployed host eggs and insecticide, are discussed. © Korean Society of Applied Entomology, Taiwan Entomological Society and Malaysian Plant Protection Society, 2012. Published by Elsevier B.V. All rights reserved.
Introduction Riptortus pedestris (Fabricius) (Hemiptera: Alydidae) is one of the most important soybean pests in Korea (Son et al., 2000; Kang et al., 2003; Lee et al., 2004). Gryon japonicum (Ashmead) (Hymenoptera: Scelionidae) and Ooencyrtus nezarae Ishii (Hymenoptera: Encyrtidae) are important egg parasitoids of R. pedestris (Paik et al., 2007). Gryon japonicum is a solitary egg parasitoid attacking only R. pedestris, whereas O. nezarae is gregarious and parasitizes the eggs of several stink bugs, including Piezodorus hybneri (Gmelin) (Hemiptera: Pentatomidae), Dolycoris baccarum (L.) (Hemiptera: Pentatomidae), and Nezara antennata Scott (Hemiptera: Pentatomidae) (Takasu and Hirose, 1985; Hirose et al., 1996; Mizutani et al., 1996; Zhang et al., 2005). Augmentation of these egg parasitoids against R. pedestris may not be economically feasible in a low-cash value crop like soybeans. Instead, augmentation of the host egg supply has been suggested as a method to increase field parasitism (Lim and Mahmoud, 2009, Alim and Lim, in press). As releases of viable host eggs would increase the field pest population, Lim and Mahmoud (2009) used refrigerated, non-viable eggs for host augmentation. Host eggs were rendered non-viable after 30 days of refrigeration, which kills eggs but leaves them suitable for parasitization by egg parasitoids, as shown in other ⁎ Corresponding author. Tel.: + 82 54 820 5510; fax: + 82 54 823 1628. E-mail address: [email protected] (U.T. Lim).
stink bugs (Orr, 1988; Kivan and Kilic, 2005; Mahmoud and Lim, 2007; Alim and Lim, 2009). While higher parasitism by G. japonicum was found in both natural and refrigerated eggs, in treated soybean fields, there were no corresponding significant reductions in stink bug populations observed; this may be because adult R. pedestris are highly mobile (Son et al., 2000; Wada et al., 2006) and thus might move into the host egg treated fields from elsewhere. One solution to this problem could be the use of insecticide in a way compatible with key natural enemies. Population density of adult R. pedestris often remains high until harvest (Lim and Mahmoud, 2009; Alim and Lim, in press) because adult bugs may invade a field, and local parasitism cannot immediately reduce adult stink bug populations. This period close to soybean harvest is the best time for insecticide application to reduce adult R. pedestris population with minimal damage to its parasitoids. This study, in which we used a cage exclusion design, tested the compatibility of combining releases of refrigerated host eggs with an insecticide application in soybean. Materials and methods Insect rearing and production of host eggs Adult R. pedestris were collected and maintained in the laboratory, as described by Alim and Lim (2009). Nymphs and adults were kept in separate acrylic cages (40 L × 40 W × 40 H cm), with mesh screens
1226-8615/$ – see front matter © Korean Society of Applied Entomology, Taiwan Entomological Society and Malaysian Plant Protection Society, 2012. Published by Elsevier B.V. All rights reserved. doi:10.1016/j.aspen.2012.03.008
300
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
in three of the sides, for ventilation, and held at 24.1 to 28.8 °C. Adult R. pedestris were provided with soybean seeds and ascorbic acid dissolved in water (2%). Nymphs were fed soybean seeds and red kidney bean plants in the cotyledon stage. Four pieces of gauze, in order that these might be used as oviposition substrates, were placed in both the upper and lower corners of the cages of adult bugs. Eggs were collected daily and held in zip-lock type plastic bags at 2.1 ± 0.7 °C. Soybean cultivation and experiment layout The experiment was carried out in a soybean field (45 × 26 m) at Songcheon, Andong, Republic of Korea. Seeds of the soybean variety Agakong were sown on 6 June 2009, with a 20 cm spacing between plants. Agakong, developed by crossing Eunhakong, a yellow-seeded sprout soybean (Glycine max L. Merr.) and KLG10084, a wild soybean with a green seed coat (Glycine soja Sieb & Zucc.) (Lee et al., 2005), is a unique soybean variety, containing three times more isoflavone than other varieties and having a lower plant canopy and smaller pod size (Kim and Lim, 2010). The experimental field was divided into 16 large plots (10 × 6 m), of which 15 were assigned randomly to the three treatments: the release of refrigerated eggs (T1), the release of refrigerated eggs with a single application of thiamethoxam (a chlorothiazol type neonicotinoid insecticide registered for the control of stink bugs on various crops in Korea [KCPA, 2007]) (T2), and the untreated control. Each treatment was replicated five times. White fabric (without insect permeable pores) was used to erect walls (height 1.1 m) that enclosed the center portion of each plot (a 3 × 2 m area) so as to minimize stink bug and parasitoids’ movement into or out of the sampled portions of the plots (= open-cage design). Chemical fertilizers or herbicides were not applied to the field, but plots were irrigated as needed.
every 6 days until 30 September, i.e., eight times during the experiment, which ended on 6 October when the soybeans were harvested. Sampling of R. pedestris Whole plant counts were taken every 6 days on three randomly selected plants in each plot, from 22 August to 3 October 2009 in order to record the occurrence of eggs, nymphs, and adults of R. pedestris in the plots. Other stink bug species were also recorded. Insecticide application When soybeans were in the R7 growth stage, five plots (Treatment T2) were sprayed with thiamethoxam, at the field recommended rate, on 22 September. Statistical analyses Data on all observations, such as R. pedestris density, other stink bug densities, and parasitism by O. nezarae and G. japonicum, were subjected to square root transformation and were analyzed using repeated measures ANOVA, using GLM procedure in SAS (SAS Institute, 1995). Tukey's studentized range honestly significant difference (HSD) test was used for further separation of least square means among the treatments on each sampling date. Results Density of Riptortus pedestris
A hundred R. pedestris eggs, which had been kept at 2.1 ± 0.7 °C for 30–35 days, were placed in each plot (in two net pouches of 2 × 2 mm mesh size, each containing 50 eggs) on 12 August 2009, before the plots were separated with fabric barriers (described above) on 19 August. After 1 week of field exposure, the refrigerated eggs were recovered on 19 August and pooled, from which 40 eggs were selected randomly; this number was placed in each of the 10 treatment plots in order to enhance the parasitoid population. Fifty-nine eggs from the pooled eggs were placed individually in micro tubes (2 ml) and held in a growth chamber (30 °C) for parasitoid emergence to estimate the initial parasitism rate. Additional refrigerated eggs (100 per plot) were placed in the field on 19 August as host sources for parasitoids. Those eggs were recollected from the field after 1 week (on 26 August) and were again re-distributed evenly over all the treatment plots in order to increase the parasitoid population. The reason for releasing only twice was to observe the multiplication of the parasitoids from initial release of refrigerated eggs, before R. pedestris became abundant.
Adult R. pedestris were found in the field from the first sampling date on 22 August through the last sampling date on 15 September, and there were no significant differences among treatments (F = 1.49, df= 14, 84, P = 0.133; Fig. 1A). Changes in the R. pedestris adult population were highly significant over the whole sampling period (F = 5.61, df= 7, 84, P b 0.001) as well as among treatments (F = 9.42, 12, df= 2, P = 0.004). There was a trend in the data for the adult population in control plots to exceed those in the treatment of refrigerated eggs (T1), and this difference was significant on 21 September (F = 4.90, df= 2, 12, P = 0.028) and 3 October (F = 4.70, df = 2, 12, P = 0.031). In the plots (T2) where pesticide was applied on 22 September, though the adult population was lowest following the application, there was no statistical significance (F = 1.60, df= 2, 12, P = 0.241). In contrast, the number of nymphs varied significantly over time (F = 3.23, df= 7, 84, P = 0.004) but did not vary among treatments (F = 0.25, df= 2, 12, P = 0.779), and there were no significant interaction effects for the nymph population size (F = 1.06, df = 14, 84, P = 0.409; Fig. 1B). The number of eggs collected was not significantly different among treatments (F = 1.17, df = 2, 12, P = 0.343), but there were significant time (F = 2.68, df= 7, 84, P = 0.015) and interaction effects (F = 1.86, df= 14, 84, P = 0.043) (Fig. 1C).
Release of unrefrigerated eggs
Egg parasitism
Unrefrigerated eggs (less than 1 day old) were deployed in each plot in order to measure the rates of natural parasitism as naturally occurring eggs were not abundant enough for analysis, especially in the beginning of the experiment. Unrefrigerated eggs of R. pedestris were obtained from our laboratory colony, and 60 eggs were placed in each plot (in two pouches of 30 each), for the first time on 19 August. From these deployed eggs, 30 were collected from each plot 3 days later (prior to the hatching of unparasitized eggs) to estimate field parasitism rates; they were held individually in micro tubes (2 ml) at 30 °C, in a growth chamber, until adult parasitoid emergence. Unrefrigerated eggs of R. pedestris were deployed in this manner
Initial parasitism in the field-deployed refrigerated eggs retrieved on 19 August was 18.6% by G. japonicum and 10.2% by O. nezarae. Parasitism by both G. japonicum and O. nezarae was also detected in batches of unrefrigerated eggs that were retrieved 3 days after their deployment in the field; their parasitism rates over time are presented in Fig. 2A (G. japonicum) and 2B (O. nezarae). Parasitism by G. japonicum was significantly higher in both treatment plots (T1 and T2) on four sampling dates (3 September F = 3.97, df= 2, 12, P = 0.048; 21 September F = 4.61, df= 2, 12, P = 0.033). Parasitism in T2 plots decreased sharply after the application of pesticide, causing parasitism by G. japonicum in T1 plots to be higher than that in T2 or untreated plots (27 September
Release of refrigerated eggs
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
301
Fig. 1. Occurrence (mean number+ SE) of R. pedestris adults (A), nymphs (B), and eggs (C) in three different treatments; release of refrigerated eggs (T1), release of refrigerated eggs with one-time spray of thiamethoxam (T2), and untreated control, in Agakong soybean. *0.01 b P ≤ 0.05 from a repeated measure of ANOVA. Arrow indicates date of thiamethoxam spray.
F = 13.64, df = 2, 12, P b 0.001; 3 October F = 7.19, df= 2, 12, P = 0.009). Parasitism rates by G. japonicum were significantly different among the treatments (F = 7.13, df= 2, 12, P = 0.009) and among sample dates
(F = 3.00, df = 7, 84, P = 0.007). However, for this parasitoid there was no significant interaction between time and treatment (F = 1.60, df= 14, 84, P = 0.095).
302
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
Fig. 2. Parasitism rate (+ SE) of released eggs of R. pedestris by G. japonicum (A) and O. nezarae (B) in three different treatments. *0.01 b P ≤ 0.05; **0.001 b P ≤ 0.01; *** P b 0.001 from a repeated measure of ANOVA. Arrow indicates date of thiamethoxam spray.
In contrast to G. japonicum, O. nezarae parasitism was very low on all sample dates. No significant differences were found in parasitism rates for O. nezarae among the treatments (F = 0.17, df = 2, 12,
P = 0.845) or for the interaction between time and treatment (F = 1.36, df= 14, 84, P = 0.193). Significant differences were found for this parasitoid among sample dates (F = 4.14, df = 7, 84, P = 0.001).
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
303
Fig. 3. Occurrence (mean number + SE) of nymph and adult populations of H. halys (A), D. baccarum (B), and Nezara spp. (C) in three different treatments. Arrow indicates date of thiamethoxam spray.
304
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
Occurrence of other stink bugs Other stink bugs found in the sampled soybean field were Halyomorpha halys (Stål) (Hemiptera: Pentatomidae), D. baccarum and Nezara spp. (Fig. 3). The density of H. halys and D. baccarum nymphs and adults fluctuated throughout the sampling period. However, after insecticide application, densities of H. halys nymphs and adults were significantly reduced in T2 plots. The greatest density of H. halys was in the untreated control plots on 27 September (F = 14.51, df= 2, 12, P = 0.001). No significant differences were found for H. halys density among treatments (F = 0.63, df= 2, 12, P = 0.551) or in the time by treatment interaction (F = 1.75, df = 14, 84, P = 0.062). However, significant differences were found for this species over sample dates (F = 8.40, df= 7, 84, P b 0.001). Nymph and adult populations of D. baccarum were not significantly different among treatments (F = 0.05, df= 2, 12, P = 0.956) or over the time period (F = 1.62, df = 7, 84, P = 0.140). However, the interaction of time and treatment was significant (F = 2.22, df= 14, 84, P = 0.013). Compared to other stink bugs, densities of the nymph and adults of Nezara spp. were higher and varied significantly among sample dates (F = 14.68, df = 7, 84, P b 0.001). However, there was no significant difference of this species’ density among treatments (F = 2.26, df = 2, 12, P = 0.147) even though the interaction of time and treatment was significant (F = 2.28, df = 14, 84, P = 0.011). Discussion We observed a significant reduction in the density of adults of R. pedestris in host egg-released plots twice during the last three sample periods, but no significant reduction was recorded in nymphal and egg densities among treatments. The insignificance of the observed numbers of eggs could be due to the counting of both parasitized and unparasitized eggs during sampling, as they could not be differentiated in the field, and it takes over 2 weeks for a parasitoid to emerge from the eggs. Also, reduction in the adult population during final samplings probably resulted from the accumulated impact on the number, as R. pedestris, as an adult, lives longer than other life stages. However, the reason for the insignificance in the number of nymphs is unknown. Increase in the release rate or the frequency of the refrigerated eggs might help to effectively reduce the different stages of bugs’ densities. Pesticide application did not significantly reduce the densities of pest bugs. The insecticide application failed to reduce the stink bug population because the bugs, due to their high mobility, could escape the effects of the pesticides (Son et al., 2000; Wada et al., 2006). Also, resistance to a pesticide could be a possible reason, although pesticide resistance in R. pedestris has not been reported to date. Therefore, more insecticide applications, or other management tactics, would be necessary to reduce the late season population of R. pedestris. Pesticide use did cause a rapid reduction in parasitism by G. japonicum in the plots treated with thiamethoxam. All and Treacy (2006) categorized thiamethoxam as an insecticide causing moderate toxicity (31–70% reduction in numbers) in parasitic wasps, but high toxicity (70% reduction in numbers) in hemipteran and neuropteran natural enemies. Lim and Mahmoud (2008) also reported that thiamethoxam spray could be harmful to Trissolcus nigripedius Nakagawa (Hymenoptera: Scelionidae). Nevertheless, lower parasitism caused by the insecticide application late in the growing season might not lead to an increase in soybean damage close to the harvest. Younger life stages of the bugs may not have strong enough stylets to penetrate maturing soybean pods, and, before they molt and become adults, the crop might have already been harvested. We found higher field parasitism in the plot where refrigerated eggs were released, as did Lim and Mahmoud (2009). Although parasitism was significantly higher in treated plots, compared to the control plot, overall parasitism was low compared to the previous study, in
which parasitism was found to reach more than 60% in the treated sites. This might be caused by a shorter period of exposure of the eggs in this study, which may have limited time that parasitoids could find the eggs. As the released host eggs were fresh ones and start to hatch in four days in field, we recollected the eggs after 3 days of the release. Another reason for lower parasitism could be the smaller number of the refrigerated eggs released. The optimum release density of refrigerated eggs for reduction of the bug population should be determined in the future. Parasitism rates by G. japonicum were higher than those of O. nezarae throughout the sampling period. Lim and Mahmoud (2009) reported increased parasitism in fields inoculated with refrigerated eggs and, likewise, found lower parasitism by O. nezarae. Interspecific competition between the parasitoids might have influenced parasitism rates. Probably, G. japonicum arrived in the field earlier than O. nezarae. A generally higher abundance of G. japonicum, compared to O. nezarae, was recorded early in the season (Mainali and Lim, 2012). As host eggs were exposed for only 3 days, there was a greater chance for G. japonicum to outcompete O. nezarae, because G. japonicum can successfully compete with O. nezarae in a host egg previously parasitized by O. nezarae for fewer than 3 days (Mizutani, 1994). The limited mobility of O. nezarae, due to caging, could be another reason for its lower parasitism rate as O. nezarae usually flies within the canopy and walks on individual plants to find host eggs (Takasu et al., 2004). Interestingly, we found a high density of Nezara spp. throughout the sampling period. Nezara antennata (Ishii, 1928) and N. viridula L. (Hokyo, 1965) are also hosts of O. nezarae. Low densities of O. nezarae, limiting the parasitism of Nezara spp. eggs might be one of the reasons for the successful adaptation of the green stink bug on Agakong soybeans. Generally, the density of this bug is low in soybean fields (Kim and Lim, 2010). Further studies on the interaction of soybean variety and Nezara spp. density should be undertaken, as this study suggests that this bug maintains a higher density in soybeans than do other soybean bugs. In conclusion, this cage exclusion test verified previous findings that the release of refrigerated eggs, in batches, into soybean fields can enhance field parasitism rates. However, one-time application of insecticide late in the growing season was found to have little effect on the density of R. pedestris in soybean fields, but it did cause significant reduction in parasitism. Acknowledgments We thank Md. Abdul Alim for the help in sampling. This study was carried out with the support of the Technology Development Program for Agriculture and Forestry, Ministry for Agriculture, Forestry and Fisheries, Republic of Korea. Bishwo P. Mainali and Sangwon Kim were supported by the 2nd Stage BK21 program of Ministry of Education, Science, and Technology, Republic of Korea. References Alim, M.A., Lim, U.T., 2009. Refrigeration of Riptortus clavatus (Hemiptera: Alydidae) eggs for the parasitization by Gryon japonicum (Hymenoptera: Scelionidae). Biocontrol Sci. Technol. 19, 315–325. Alim, M.A., Lim, U.T., 2011. Refrigerated eggs of Riptortus pedestris (Hemiptera: Alydidae) added to aggregation pheromone traps increase field parasitism in soybean. J. Econ. Entomol. 104, 1833–1839. All, J.N., Treacy, M.F., 2006. Use and management of insecticides, acaricides, and transgenic crops. Entomol. Soc. Am.Lanham, MD, USA. Hirose, Y., Takasu, K., Takagi, M., 1996. Egg parasitoids of phytophagous bugs in soybean: mobile natural enemies as naturally occurring biological control agents of mobile pests. Biol. Control 7, 84–94. Hokyo, N., 1965. Interspecific relations among egg parasites of Nezara viridula L., with special reference to Asolcus mitsukurii Ashmead and Telenomus nakagawai Watanabe. Nanki-seibutsu 7, 1–6 (in Japanese). Kang, C.H., Huh, H.S., Park, C.G., 2003. Review on true bugs infesting tree fruit, upland crops, and weeds in Korea. Korean J. Appl. Entomol. 42, 269–277. KCPA, 2007. Agrochemicals use guide book. Korea Crop Protection Association, Seoul, Korea.
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305 Kim, S., Lim, U.T., 2010. Seasonal occurrence pattern and within-plant egg distribution of bean bug, Riptortus pedestris (Fabricius) (Hemiptera: Alydidae), and its egg parasitoids in soybean fields. Appl. Entomol. Zool. 45, 457–464. Kivan, M., Kilic, N., 2005. Effects of storage at low-temperature of various heteropteran host eggs on the egg parasitoid, Trissolcus semistriatus. BioControl 50, 589–600. Lee, G.H., Paik, C.H., Choi, M.Y., Oh, Y.J., Kim, D.H., Na, S.Y., 2004. Seasonal occurrence, soybean damage and control efficacy of bean bug, Riptortus clavatus Thunberg (Hemiptera: Alydidae) at soybean field in Honam Province. Korean J. Appl. Entomol. 43, 249–255. Lee, J.D., Jeong, Y.S., Grover Shannon, J., Park, S.K., Choung, M.G., Hwang, Y.H., 2005. Agronomic characteristics of small-seeded RILs derived from Eunhakong (Glycine max) × KLG10048 (G. soja). Korean J. Breed. 37, 288–294. Lim, U.T., Mahmoud, A.M.A., 2008. Ecotoxicological effect of fenitrothion on Trissolcus nigripedius (Hymenoptera: Scelionidae) an egg parasitoid of Dolycoris baccarum (Hemiptera: Pentatomidae). J. Asia Pac. Entomol. 11, 207–210. Lim, U.T., Mahmoud, A.M.A., 2009. Inoculation of refrigerated non-viable eggs of Riptortus clavatus (Heteroptera: Alydidae) to enhance parasitism by egg parasitoid in soybean field. Appl. Entomol. Zool. 44, 37–45. Mahmoud, A.M.A., Lim, U.T., 2007. Evaluation of cold-stored eggs of Dolycoris baccarum (Hemiptera: Pentatomidae) for parasitization by Trissolcus nigripedius (Hymenoptera: Scelionidae). Biol. Control 43, 287–293. Mainali, B.P., Lim, U.T., 2012. Annual pattern of occurrence of Riptortus pedestris (Hemiptera: Alydidae) and its egg parasitoids Ooencyrtus nezarae Ishii and Gryon japonicum (Ashmead) in Andong, Korea. Crop Prot. 36, 37–42.
305
Mizutani, N., Hirose, Y., Higuchi, H., Wada, T., 1996. Seasonal abundance of Ooencyrtus nezarae Ishii (Hymenoptera: Encyrtidae), an egg parasitoid of phytophagous bugs, in summer soybean fields. Jpn. J. Appl. Entomol. Zool. 40, 199–204. Mizutani, N., 1994. Interspecific larval competition among three egg parasitoid species on the host, Riptortus clavatus (Thunberg) (Heteroptera: Alydidae). Proc. Assoc. Plant Prot. Kyushu 40, 106–110. Orr, D.B., 1988. Scelionid wasps as biological control agents: a review. Fla. Entomol. 71, 506–528. Paik, C.H., Lee, G.H., Choi, M.Y., Seo, H.Y., Kim, D.H., La, S.Y., Park, C.G., 2007. Report on two egg parasitoids of Riptortus clavatus (Thunberg) (Heteroptera: Alydidae) on soybean. Korean J. Appl. Entomol. 46, 281–286. SAS Institute, 1995. SAS for Windows. Release 6, 11 ed. SAS Institute, Cary, NC, USA. Son, C.K., Park, S.G., Hwang, Y.H., Choi, B.S., 2000. Field occurrence of stink bug and its damage in soybean. Korean J. Crop Sci. 45, 405–410. Takasu, K., Hirose, Y., 1985. Seasonal egg parasitism of phytophagous stink bugs in a soybean field in Fukuoka. Proc. Assoc. Plant Prot. Kyushu 31, 127–131. Takasu, K., Takano, S.I., Mizutani, N., Wada, T., 2004. Flight orientation behavior of Ooencyrtus nezarae (Hymenoptera: Encyrtidae), an egg parasitoid of phytophagous bugs in soybean. Entomol Sci. 7, 201–206. Wada, T., Endo, N., Takahashi, M., 2006. Reducing seed damage by soybean bugs by growing small-seeded soybeans and delaying sowing time. J. Crop Prot. 25, 726–731. Zhang, Y.Z., Li, W., Huang, D.W., 2005. A taxonomic study of Chinese species of Ooencyrtus (Insecta: Hymenoptera: Encyrtidae). Zool. Stud. 44, 347–360.
Contents lists available at SciVerse ScienceDirect
Journal of Asia-Pacific Entomology journal homepage: www.elsevier.com/locate/jape
Effects of combining releases of non-viable host eggs with insecticide application on Riptortus pedestris population and its egg parasitoids Bishwo P. Mainali a, Sangwon Kim b, Un Taek Lim a,⁎ a b
School of Bioresource Sciences, Andong National University, Andong 760–749, Republic of Korea Dongbu Ceres Co. Ltd., 135 Dongsan, Nonsan, Republic of Korea
a r t i c l e
i n f o
Article history: Received 8 December 2011 Revised 24 March 2012 Accepted 31 March 2012 Keywords: Ooencyrtus nezarae Gryon japonicum Cold storage Parasitism Thiamethoxam Alydid
a b s t r a c t Our previous study demonstrated that the release of refrigerated non-viable eggs of Riptortus pedestris (Fabricius) (Hemiptera: Alydidae) enhanced parasitism rates in soybean fields but did not result in the reduction of R. pedestris populations. This study was further conducted using an open-cage exclusion design in a soybean field in order to evaluate the compatibility of combining releases of non-viable host eggs with a single pre-harvest application of insecticide for the control of R. pedestris. Refrigerated eggs of R. pedestris were released twice in treatment plots, and fresh (b 1 day old) eggs of R. pedestris were deployed in all experimental arenas, every 6 days, for host resource and measurement of field parasitism. The releases of host eggs did not reduce the number of R. pedestris in any life stage except the adult stage on two sampling dates. However, parasitism by Gryon japonicum (Ashmead) (Hymenoptera: Scelionidae) was higher in treated plots (9–25%) than in the control plots (1–9%). Statistical significant reduction was not found in the pest population, but parasitism rates significantly increased. Pesticide application did not reduce the bug population but did affect the parasitoids population. Pest management tactics, using both artificially deployed host eggs and insecticide, are discussed. © Korean Society of Applied Entomology, Taiwan Entomological Society and Malaysian Plant Protection Society, 2012. Published by Elsevier B.V. All rights reserved.
Introduction Riptortus pedestris (Fabricius) (Hemiptera: Alydidae) is one of the most important soybean pests in Korea (Son et al., 2000; Kang et al., 2003; Lee et al., 2004). Gryon japonicum (Ashmead) (Hymenoptera: Scelionidae) and Ooencyrtus nezarae Ishii (Hymenoptera: Encyrtidae) are important egg parasitoids of R. pedestris (Paik et al., 2007). Gryon japonicum is a solitary egg parasitoid attacking only R. pedestris, whereas O. nezarae is gregarious and parasitizes the eggs of several stink bugs, including Piezodorus hybneri (Gmelin) (Hemiptera: Pentatomidae), Dolycoris baccarum (L.) (Hemiptera: Pentatomidae), and Nezara antennata Scott (Hemiptera: Pentatomidae) (Takasu and Hirose, 1985; Hirose et al., 1996; Mizutani et al., 1996; Zhang et al., 2005). Augmentation of these egg parasitoids against R. pedestris may not be economically feasible in a low-cash value crop like soybeans. Instead, augmentation of the host egg supply has been suggested as a method to increase field parasitism (Lim and Mahmoud, 2009, Alim and Lim, in press). As releases of viable host eggs would increase the field pest population, Lim and Mahmoud (2009) used refrigerated, non-viable eggs for host augmentation. Host eggs were rendered non-viable after 30 days of refrigeration, which kills eggs but leaves them suitable for parasitization by egg parasitoids, as shown in other ⁎ Corresponding author. Tel.: + 82 54 820 5510; fax: + 82 54 823 1628. E-mail address: [email protected] (U.T. Lim).
stink bugs (Orr, 1988; Kivan and Kilic, 2005; Mahmoud and Lim, 2007; Alim and Lim, 2009). While higher parasitism by G. japonicum was found in both natural and refrigerated eggs, in treated soybean fields, there were no corresponding significant reductions in stink bug populations observed; this may be because adult R. pedestris are highly mobile (Son et al., 2000; Wada et al., 2006) and thus might move into the host egg treated fields from elsewhere. One solution to this problem could be the use of insecticide in a way compatible with key natural enemies. Population density of adult R. pedestris often remains high until harvest (Lim and Mahmoud, 2009; Alim and Lim, in press) because adult bugs may invade a field, and local parasitism cannot immediately reduce adult stink bug populations. This period close to soybean harvest is the best time for insecticide application to reduce adult R. pedestris population with minimal damage to its parasitoids. This study, in which we used a cage exclusion design, tested the compatibility of combining releases of refrigerated host eggs with an insecticide application in soybean. Materials and methods Insect rearing and production of host eggs Adult R. pedestris were collected and maintained in the laboratory, as described by Alim and Lim (2009). Nymphs and adults were kept in separate acrylic cages (40 L × 40 W × 40 H cm), with mesh screens
1226-8615/$ – see front matter © Korean Society of Applied Entomology, Taiwan Entomological Society and Malaysian Plant Protection Society, 2012. Published by Elsevier B.V. All rights reserved. doi:10.1016/j.aspen.2012.03.008
300
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
in three of the sides, for ventilation, and held at 24.1 to 28.8 °C. Adult R. pedestris were provided with soybean seeds and ascorbic acid dissolved in water (2%). Nymphs were fed soybean seeds and red kidney bean plants in the cotyledon stage. Four pieces of gauze, in order that these might be used as oviposition substrates, were placed in both the upper and lower corners of the cages of adult bugs. Eggs were collected daily and held in zip-lock type plastic bags at 2.1 ± 0.7 °C. Soybean cultivation and experiment layout The experiment was carried out in a soybean field (45 × 26 m) at Songcheon, Andong, Republic of Korea. Seeds of the soybean variety Agakong were sown on 6 June 2009, with a 20 cm spacing between plants. Agakong, developed by crossing Eunhakong, a yellow-seeded sprout soybean (Glycine max L. Merr.) and KLG10084, a wild soybean with a green seed coat (Glycine soja Sieb & Zucc.) (Lee et al., 2005), is a unique soybean variety, containing three times more isoflavone than other varieties and having a lower plant canopy and smaller pod size (Kim and Lim, 2010). The experimental field was divided into 16 large plots (10 × 6 m), of which 15 were assigned randomly to the three treatments: the release of refrigerated eggs (T1), the release of refrigerated eggs with a single application of thiamethoxam (a chlorothiazol type neonicotinoid insecticide registered for the control of stink bugs on various crops in Korea [KCPA, 2007]) (T2), and the untreated control. Each treatment was replicated five times. White fabric (without insect permeable pores) was used to erect walls (height 1.1 m) that enclosed the center portion of each plot (a 3 × 2 m area) so as to minimize stink bug and parasitoids’ movement into or out of the sampled portions of the plots (= open-cage design). Chemical fertilizers or herbicides were not applied to the field, but plots were irrigated as needed.
every 6 days until 30 September, i.e., eight times during the experiment, which ended on 6 October when the soybeans were harvested. Sampling of R. pedestris Whole plant counts were taken every 6 days on three randomly selected plants in each plot, from 22 August to 3 October 2009 in order to record the occurrence of eggs, nymphs, and adults of R. pedestris in the plots. Other stink bug species were also recorded. Insecticide application When soybeans were in the R7 growth stage, five plots (Treatment T2) were sprayed with thiamethoxam, at the field recommended rate, on 22 September. Statistical analyses Data on all observations, such as R. pedestris density, other stink bug densities, and parasitism by O. nezarae and G. japonicum, were subjected to square root transformation and were analyzed using repeated measures ANOVA, using GLM procedure in SAS (SAS Institute, 1995). Tukey's studentized range honestly significant difference (HSD) test was used for further separation of least square means among the treatments on each sampling date. Results Density of Riptortus pedestris
A hundred R. pedestris eggs, which had been kept at 2.1 ± 0.7 °C for 30–35 days, were placed in each plot (in two net pouches of 2 × 2 mm mesh size, each containing 50 eggs) on 12 August 2009, before the plots were separated with fabric barriers (described above) on 19 August. After 1 week of field exposure, the refrigerated eggs were recovered on 19 August and pooled, from which 40 eggs were selected randomly; this number was placed in each of the 10 treatment plots in order to enhance the parasitoid population. Fifty-nine eggs from the pooled eggs were placed individually in micro tubes (2 ml) and held in a growth chamber (30 °C) for parasitoid emergence to estimate the initial parasitism rate. Additional refrigerated eggs (100 per plot) were placed in the field on 19 August as host sources for parasitoids. Those eggs were recollected from the field after 1 week (on 26 August) and were again re-distributed evenly over all the treatment plots in order to increase the parasitoid population. The reason for releasing only twice was to observe the multiplication of the parasitoids from initial release of refrigerated eggs, before R. pedestris became abundant.
Adult R. pedestris were found in the field from the first sampling date on 22 August through the last sampling date on 15 September, and there were no significant differences among treatments (F = 1.49, df= 14, 84, P = 0.133; Fig. 1A). Changes in the R. pedestris adult population were highly significant over the whole sampling period (F = 5.61, df= 7, 84, P b 0.001) as well as among treatments (F = 9.42, 12, df= 2, P = 0.004). There was a trend in the data for the adult population in control plots to exceed those in the treatment of refrigerated eggs (T1), and this difference was significant on 21 September (F = 4.90, df= 2, 12, P = 0.028) and 3 October (F = 4.70, df = 2, 12, P = 0.031). In the plots (T2) where pesticide was applied on 22 September, though the adult population was lowest following the application, there was no statistical significance (F = 1.60, df= 2, 12, P = 0.241). In contrast, the number of nymphs varied significantly over time (F = 3.23, df= 7, 84, P = 0.004) but did not vary among treatments (F = 0.25, df= 2, 12, P = 0.779), and there were no significant interaction effects for the nymph population size (F = 1.06, df = 14, 84, P = 0.409; Fig. 1B). The number of eggs collected was not significantly different among treatments (F = 1.17, df = 2, 12, P = 0.343), but there were significant time (F = 2.68, df= 7, 84, P = 0.015) and interaction effects (F = 1.86, df= 14, 84, P = 0.043) (Fig. 1C).
Release of unrefrigerated eggs
Egg parasitism
Unrefrigerated eggs (less than 1 day old) were deployed in each plot in order to measure the rates of natural parasitism as naturally occurring eggs were not abundant enough for analysis, especially in the beginning of the experiment. Unrefrigerated eggs of R. pedestris were obtained from our laboratory colony, and 60 eggs were placed in each plot (in two pouches of 30 each), for the first time on 19 August. From these deployed eggs, 30 were collected from each plot 3 days later (prior to the hatching of unparasitized eggs) to estimate field parasitism rates; they were held individually in micro tubes (2 ml) at 30 °C, in a growth chamber, until adult parasitoid emergence. Unrefrigerated eggs of R. pedestris were deployed in this manner
Initial parasitism in the field-deployed refrigerated eggs retrieved on 19 August was 18.6% by G. japonicum and 10.2% by O. nezarae. Parasitism by both G. japonicum and O. nezarae was also detected in batches of unrefrigerated eggs that were retrieved 3 days after their deployment in the field; their parasitism rates over time are presented in Fig. 2A (G. japonicum) and 2B (O. nezarae). Parasitism by G. japonicum was significantly higher in both treatment plots (T1 and T2) on four sampling dates (3 September F = 3.97, df= 2, 12, P = 0.048; 21 September F = 4.61, df= 2, 12, P = 0.033). Parasitism in T2 plots decreased sharply after the application of pesticide, causing parasitism by G. japonicum in T1 plots to be higher than that in T2 or untreated plots (27 September
Release of refrigerated eggs
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
301
Fig. 1. Occurrence (mean number+ SE) of R. pedestris adults (A), nymphs (B), and eggs (C) in three different treatments; release of refrigerated eggs (T1), release of refrigerated eggs with one-time spray of thiamethoxam (T2), and untreated control, in Agakong soybean. *0.01 b P ≤ 0.05 from a repeated measure of ANOVA. Arrow indicates date of thiamethoxam spray.
F = 13.64, df = 2, 12, P b 0.001; 3 October F = 7.19, df= 2, 12, P = 0.009). Parasitism rates by G. japonicum were significantly different among the treatments (F = 7.13, df= 2, 12, P = 0.009) and among sample dates
(F = 3.00, df = 7, 84, P = 0.007). However, for this parasitoid there was no significant interaction between time and treatment (F = 1.60, df= 14, 84, P = 0.095).
302
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
Fig. 2. Parasitism rate (+ SE) of released eggs of R. pedestris by G. japonicum (A) and O. nezarae (B) in three different treatments. *0.01 b P ≤ 0.05; **0.001 b P ≤ 0.01; *** P b 0.001 from a repeated measure of ANOVA. Arrow indicates date of thiamethoxam spray.
In contrast to G. japonicum, O. nezarae parasitism was very low on all sample dates. No significant differences were found in parasitism rates for O. nezarae among the treatments (F = 0.17, df = 2, 12,
P = 0.845) or for the interaction between time and treatment (F = 1.36, df= 14, 84, P = 0.193). Significant differences were found for this parasitoid among sample dates (F = 4.14, df = 7, 84, P = 0.001).
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
303
Fig. 3. Occurrence (mean number + SE) of nymph and adult populations of H. halys (A), D. baccarum (B), and Nezara spp. (C) in three different treatments. Arrow indicates date of thiamethoxam spray.
304
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305
Occurrence of other stink bugs Other stink bugs found in the sampled soybean field were Halyomorpha halys (Stål) (Hemiptera: Pentatomidae), D. baccarum and Nezara spp. (Fig. 3). The density of H. halys and D. baccarum nymphs and adults fluctuated throughout the sampling period. However, after insecticide application, densities of H. halys nymphs and adults were significantly reduced in T2 plots. The greatest density of H. halys was in the untreated control plots on 27 September (F = 14.51, df= 2, 12, P = 0.001). No significant differences were found for H. halys density among treatments (F = 0.63, df= 2, 12, P = 0.551) or in the time by treatment interaction (F = 1.75, df = 14, 84, P = 0.062). However, significant differences were found for this species over sample dates (F = 8.40, df= 7, 84, P b 0.001). Nymph and adult populations of D. baccarum were not significantly different among treatments (F = 0.05, df= 2, 12, P = 0.956) or over the time period (F = 1.62, df = 7, 84, P = 0.140). However, the interaction of time and treatment was significant (F = 2.22, df= 14, 84, P = 0.013). Compared to other stink bugs, densities of the nymph and adults of Nezara spp. were higher and varied significantly among sample dates (F = 14.68, df = 7, 84, P b 0.001). However, there was no significant difference of this species’ density among treatments (F = 2.26, df = 2, 12, P = 0.147) even though the interaction of time and treatment was significant (F = 2.28, df = 14, 84, P = 0.011). Discussion We observed a significant reduction in the density of adults of R. pedestris in host egg-released plots twice during the last three sample periods, but no significant reduction was recorded in nymphal and egg densities among treatments. The insignificance of the observed numbers of eggs could be due to the counting of both parasitized and unparasitized eggs during sampling, as they could not be differentiated in the field, and it takes over 2 weeks for a parasitoid to emerge from the eggs. Also, reduction in the adult population during final samplings probably resulted from the accumulated impact on the number, as R. pedestris, as an adult, lives longer than other life stages. However, the reason for the insignificance in the number of nymphs is unknown. Increase in the release rate or the frequency of the refrigerated eggs might help to effectively reduce the different stages of bugs’ densities. Pesticide application did not significantly reduce the densities of pest bugs. The insecticide application failed to reduce the stink bug population because the bugs, due to their high mobility, could escape the effects of the pesticides (Son et al., 2000; Wada et al., 2006). Also, resistance to a pesticide could be a possible reason, although pesticide resistance in R. pedestris has not been reported to date. Therefore, more insecticide applications, or other management tactics, would be necessary to reduce the late season population of R. pedestris. Pesticide use did cause a rapid reduction in parasitism by G. japonicum in the plots treated with thiamethoxam. All and Treacy (2006) categorized thiamethoxam as an insecticide causing moderate toxicity (31–70% reduction in numbers) in parasitic wasps, but high toxicity (70% reduction in numbers) in hemipteran and neuropteran natural enemies. Lim and Mahmoud (2008) also reported that thiamethoxam spray could be harmful to Trissolcus nigripedius Nakagawa (Hymenoptera: Scelionidae). Nevertheless, lower parasitism caused by the insecticide application late in the growing season might not lead to an increase in soybean damage close to the harvest. Younger life stages of the bugs may not have strong enough stylets to penetrate maturing soybean pods, and, before they molt and become adults, the crop might have already been harvested. We found higher field parasitism in the plot where refrigerated eggs were released, as did Lim and Mahmoud (2009). Although parasitism was significantly higher in treated plots, compared to the control plot, overall parasitism was low compared to the previous study, in
which parasitism was found to reach more than 60% in the treated sites. This might be caused by a shorter period of exposure of the eggs in this study, which may have limited time that parasitoids could find the eggs. As the released host eggs were fresh ones and start to hatch in four days in field, we recollected the eggs after 3 days of the release. Another reason for lower parasitism could be the smaller number of the refrigerated eggs released. The optimum release density of refrigerated eggs for reduction of the bug population should be determined in the future. Parasitism rates by G. japonicum were higher than those of O. nezarae throughout the sampling period. Lim and Mahmoud (2009) reported increased parasitism in fields inoculated with refrigerated eggs and, likewise, found lower parasitism by O. nezarae. Interspecific competition between the parasitoids might have influenced parasitism rates. Probably, G. japonicum arrived in the field earlier than O. nezarae. A generally higher abundance of G. japonicum, compared to O. nezarae, was recorded early in the season (Mainali and Lim, 2012). As host eggs were exposed for only 3 days, there was a greater chance for G. japonicum to outcompete O. nezarae, because G. japonicum can successfully compete with O. nezarae in a host egg previously parasitized by O. nezarae for fewer than 3 days (Mizutani, 1994). The limited mobility of O. nezarae, due to caging, could be another reason for its lower parasitism rate as O. nezarae usually flies within the canopy and walks on individual plants to find host eggs (Takasu et al., 2004). Interestingly, we found a high density of Nezara spp. throughout the sampling period. Nezara antennata (Ishii, 1928) and N. viridula L. (Hokyo, 1965) are also hosts of O. nezarae. Low densities of O. nezarae, limiting the parasitism of Nezara spp. eggs might be one of the reasons for the successful adaptation of the green stink bug on Agakong soybeans. Generally, the density of this bug is low in soybean fields (Kim and Lim, 2010). Further studies on the interaction of soybean variety and Nezara spp. density should be undertaken, as this study suggests that this bug maintains a higher density in soybeans than do other soybean bugs. In conclusion, this cage exclusion test verified previous findings that the release of refrigerated eggs, in batches, into soybean fields can enhance field parasitism rates. However, one-time application of insecticide late in the growing season was found to have little effect on the density of R. pedestris in soybean fields, but it did cause significant reduction in parasitism. Acknowledgments We thank Md. Abdul Alim for the help in sampling. This study was carried out with the support of the Technology Development Program for Agriculture and Forestry, Ministry for Agriculture, Forestry and Fisheries, Republic of Korea. Bishwo P. Mainali and Sangwon Kim were supported by the 2nd Stage BK21 program of Ministry of Education, Science, and Technology, Republic of Korea. References Alim, M.A., Lim, U.T., 2009. Refrigeration of Riptortus clavatus (Hemiptera: Alydidae) eggs for the parasitization by Gryon japonicum (Hymenoptera: Scelionidae). Biocontrol Sci. Technol. 19, 315–325. Alim, M.A., Lim, U.T., 2011. Refrigerated eggs of Riptortus pedestris (Hemiptera: Alydidae) added to aggregation pheromone traps increase field parasitism in soybean. J. Econ. Entomol. 104, 1833–1839. All, J.N., Treacy, M.F., 2006. Use and management of insecticides, acaricides, and transgenic crops. Entomol. Soc. Am.Lanham, MD, USA. Hirose, Y., Takasu, K., Takagi, M., 1996. Egg parasitoids of phytophagous bugs in soybean: mobile natural enemies as naturally occurring biological control agents of mobile pests. Biol. Control 7, 84–94. Hokyo, N., 1965. Interspecific relations among egg parasites of Nezara viridula L., with special reference to Asolcus mitsukurii Ashmead and Telenomus nakagawai Watanabe. Nanki-seibutsu 7, 1–6 (in Japanese). Kang, C.H., Huh, H.S., Park, C.G., 2003. Review on true bugs infesting tree fruit, upland crops, and weeds in Korea. Korean J. Appl. Entomol. 42, 269–277. KCPA, 2007. Agrochemicals use guide book. Korea Crop Protection Association, Seoul, Korea.
B.P. Mainali et al. / Journal of Asia-Pacific Entomology 15 (2012) 299–305 Kim, S., Lim, U.T., 2010. Seasonal occurrence pattern and within-plant egg distribution of bean bug, Riptortus pedestris (Fabricius) (Hemiptera: Alydidae), and its egg parasitoids in soybean fields. Appl. Entomol. Zool. 45, 457–464. Kivan, M., Kilic, N., 2005. Effects of storage at low-temperature of various heteropteran host eggs on the egg parasitoid, Trissolcus semistriatus. BioControl 50, 589–600. Lee, G.H., Paik, C.H., Choi, M.Y., Oh, Y.J., Kim, D.H., Na, S.Y., 2004. Seasonal occurrence, soybean damage and control efficacy of bean bug, Riptortus clavatus Thunberg (Hemiptera: Alydidae) at soybean field in Honam Province. Korean J. Appl. Entomol. 43, 249–255. Lee, J.D., Jeong, Y.S., Grover Shannon, J., Park, S.K., Choung, M.G., Hwang, Y.H., 2005. Agronomic characteristics of small-seeded RILs derived from Eunhakong (Glycine max) × KLG10048 (G. soja). Korean J. Breed. 37, 288–294. Lim, U.T., Mahmoud, A.M.A., 2008. Ecotoxicological effect of fenitrothion on Trissolcus nigripedius (Hymenoptera: Scelionidae) an egg parasitoid of Dolycoris baccarum (Hemiptera: Pentatomidae). J. Asia Pac. Entomol. 11, 207–210. Lim, U.T., Mahmoud, A.M.A., 2009. Inoculation of refrigerated non-viable eggs of Riptortus clavatus (Heteroptera: Alydidae) to enhance parasitism by egg parasitoid in soybean field. Appl. Entomol. Zool. 44, 37–45. Mahmoud, A.M.A., Lim, U.T., 2007. Evaluation of cold-stored eggs of Dolycoris baccarum (Hemiptera: Pentatomidae) for parasitization by Trissolcus nigripedius (Hymenoptera: Scelionidae). Biol. Control 43, 287–293. Mainali, B.P., Lim, U.T., 2012. Annual pattern of occurrence of Riptortus pedestris (Hemiptera: Alydidae) and its egg parasitoids Ooencyrtus nezarae Ishii and Gryon japonicum (Ashmead) in Andong, Korea. Crop Prot. 36, 37–42.
305
Mizutani, N., Hirose, Y., Higuchi, H., Wada, T., 1996. Seasonal abundance of Ooencyrtus nezarae Ishii (Hymenoptera: Encyrtidae), an egg parasitoid of phytophagous bugs, in summer soybean fields. Jpn. J. Appl. Entomol. Zool. 40, 199–204. Mizutani, N., 1994. Interspecific larval competition among three egg parasitoid species on the host, Riptortus clavatus (Thunberg) (Heteroptera: Alydidae). Proc. Assoc. Plant Prot. Kyushu 40, 106–110. Orr, D.B., 1988. Scelionid wasps as biological control agents: a review. Fla. Entomol. 71, 506–528. Paik, C.H., Lee, G.H., Choi, M.Y., Seo, H.Y., Kim, D.H., La, S.Y., Park, C.G., 2007. Report on two egg parasitoids of Riptortus clavatus (Thunberg) (Heteroptera: Alydidae) on soybean. Korean J. Appl. Entomol. 46, 281–286. SAS Institute, 1995. SAS for Windows. Release 6, 11 ed. SAS Institute, Cary, NC, USA. Son, C.K., Park, S.G., Hwang, Y.H., Choi, B.S., 2000. Field occurrence of stink bug and its damage in soybean. Korean J. Crop Sci. 45, 405–410. Takasu, K., Hirose, Y., 1985. Seasonal egg parasitism of phytophagous stink bugs in a soybean field in Fukuoka. Proc. Assoc. Plant Prot. Kyushu 31, 127–131. Takasu, K., Takano, S.I., Mizutani, N., Wada, T., 2004. Flight orientation behavior of Ooencyrtus nezarae (Hymenoptera: Encyrtidae), an egg parasitoid of phytophagous bugs in soybean. Entomol Sci. 7, 201–206. Wada, T., Endo, N., Takahashi, M., 2006. Reducing seed damage by soybean bugs by growing small-seeded soybeans and delaying sowing time. J. Crop Prot. 25, 726–731. Zhang, Y.Z., Li, W., Huang, D.W., 2005. A taxonomic study of Chinese species of Ooencyrtus (Insecta: Hymenoptera: Encyrtidae). Zool. Stud. 44, 347–360.