VOL. 34, NO. 2 SOUTHWESTERN ENTOMOLOGIST JUN. 2009 Efficacy of Entomopathogenic Fungi in Suppressing Pecan Weevil, Curculio caryae (Coleoptera: Curculionidae), in Commercial Pecan Orchards David I. Shapiro-Ilan1, Ted E. Cottrell1, Wayne A. Gardner2, Robert W. Behle3, Bill Ree4, and Marvin K. Harris4 Abstract. The pecan weevil, Curculio caryae (Horn), is a key pest of pecans, Carya illinoinensis (Wangenh.) K. Koch. The entomopathogenic fungi Beauveria bassiana (Balsamo) Vuillemin and Metarhizium anisopliae (Metschnikoff) Sorokin are pathogenic to and are being developed as microbial control agents for pecan weevil. One approach to suppressing pest populations and the resultant damage might be to apply entomopathogenic fungi when adult pecan weevils are emerging from the soil. Here we report the efficacy of B. bassiana (GHA strain) and M. anisopliae (F52 strain) applied to trees in orchards at three locations: Byron, GA, Fort Valley, GA, and Comanche, TX. At Fort Valley, treatments included B. bassiana as an oil-based spray with a UV-protective screen applied to the trunk, M. anisopliae applied as an impregnated fiber band stapled onto the trunk, and a nontreated check. At Byron, GA, we compared the B. bassiana trunk treatment to a nontreated check. Treatments at the Texas location were the B. bassiana trunk application, M. anisopliae applied as a trunk band and as a soil drench, and a nontreated check. At each location, weevils were trapped and transported to the laboratory for 15 to 17 days post-treatment to record mortality and mycosis. At both Georgia locations, B. bassiana caused 80% mortality and mycosis, which was significantly greater than mortality observed in the check (33%); mortality and mycosis in the M. anisopliae treatment at Fort Valley did not differ from that observed in the check. In Texas, due to insufficient replication in plots, statistical comparison among treatments was not possible. However, mean percentages of mortality of pecan weevils after 7 and 14 days were 38 and 55% in the check, 75 and 88% in the B. bassiana-treated plots, and 57 and 75% in the M. anisopliae- treated plots. These results indicate potential for B. bassiana trunk sprays to suppress adult pecan weevil. Future research is needed to determine if the approach contributes to economic levels of crop protection. __________________ 1USDA-ARS, SE Fruit and Nut Tree Research Laboratory, Byron, GA 31008 2Department of Entomology, University of Georgia, Griffin Campus, 1109 Experiment Street, Griffin, GA 30223 3USDA-ARS-NCAUR, Peoria, IL 61604 4Department of Entomology, Texas A&M University, College Station, TX 77843 This article reports the results of research only. Mention of a proprietary product does not constitute an endorsement or recommendation for its use by USDA. 111 Introduction The pecan weevil, Curculio caryae (Horn), is a key insect pest of pecans, Carya illinoinensis (Wangenh.) K. Koch, throughout the southeastern US as well as portions of Texas and Oklahoma (Payne and Dutcher 1985). The insects have a 2- or 3-year life cycle (Harris 1985). Adult weevils emerge from soil in late July-August to feed on and oviposit in developing nuts. Once larval development is completed within the nut, 4th instars drop to the soil and burrow to a depth of 8-25 cm, and form a soil-cell to overwinter. The following fall approximately 90% of the larvae pupate and spend the next 9 months in the soil as adults before emerging. The remaining 10% of the population spend 2 years in the soil as larvae and emerge as adults in the 3rd year. Current recommendations for controlling pecan weevil consist mainly of above-ground applications of chemical insecticides (e.g., carbaryl) targeting adults in the canopy (Harris 1999, Hudson et al. 2006). Application of chemical insecticides is recommended every 7-10 days during peak emergence of pecan weevils (Ree et al. 2005, Hudson et al. 2006). Because of problems associated with resurgence of aphids and mites that often result from chemical applications (Dutcher and Payne 1985), as well as other environmental and regulatory concerns, developing alternative control strategies is desirable. Microbial pesticides such as the entomopathogenic fungi Beauveria bassiana (Balsamo) Vuillemin and Metarhizium anisopliae (Metschnikoff) Sorokin are potential alternatives (Shapiro- Ilan 2003, Shapiro-Ilan et al. 2008). Both the larval and adult stages of pecan weevil are susceptible to infection by these agents (Tedders et al. 1973; Gottwald and Tedders 1983; Harrison et al. 1993; Shapiro-Ilan et al. 2003, 2008, 2009). Prior research indicated that many emerging pecan weevils either crawl or fly to the trunk (Raney and Eikenbary 1968, Cottrell and Wood 2008). By exploiting this behavior, significant mortality may be obtained by applying the fungus to soil surrounding the trunk, or directly to the trunk, thereby targeting the insects before they enter the canopy to feed and oviposit. Indeed, recently, Shapiro-Ilan et al. (2008) reported that suppression of pecan weevil was affected by the method of applying B. bassiana; spraying the fungus directly on the trunk resulted in greater mortality of pecan weevil relative to direct application to soil. Additionally, another approach to applying fungus to the trunk, i.e., wrapping non-woven fiber bands impregnated with M. anisopliae around the trunk, caused significant mortality of emerging pecan weevils (Shapiro-Ilan et al. 2009). The objective of this study was to investigate the potential of applications of these fungi to suppress pecan weevil in commercial pecan orchards. We applied entomopathogenic fungi in pecan orchards at three weevil-infested locations: Byron, GA, Fort Valley, GA, and Comanche, TX. Materials and Methods ‘Stewart’, ‘Frotcher’, and ‘Van Demon’ varieties of pecan trees >150 years old in loamy-sand soil were used at Byron, GA. The Fort Valley, GA, orchard consisted of loamy-sand soil with ‘Stuart’ and ‘Schley’ trees approximately 60 years old. The Texas location was approximately 16 km north of Comanche, and consisted of native variety trees in a fine sandy-loam soil; trees varied in age and size (all non-pecan tree species had been removed 30 years earlier.) 112 Beauveria bassiana (GHA strain), i.e., Botanigard®, used in all field experiments was obtained from Emerald BioAgriculture Corporation (Butte, MT) as an emulsifiable oil formulation containing 2 x 1013 conidia per 946-ml container. This strain (GHA) has been labeled for use in controlling pecan weevil. Metarhizium anisopliae (F52 strain), formulated into fiber bands, was produced at the USDA- ARS Research Station in Stoneville, MS, based on procedures described by Hajek et al. (2006), and refrigerated while shipped to Byron, GA. M. anisopliae conidia for application to soil were produced through a biphasic system (liquid blastospore suspension poured into sterile rice medium in bags) based on the method of Leland et al. (2005). All fungal material was stored at approximately 4oC and used within 2 months of receipt. Before application, viability of conidia was verified on agar as described by Goettel and Inglis (1997), and >80% viability was deemed acceptable. At the two orchards in Georgia, the experiments were in randomized complete block designs with six blocks (tree rows) containing each treatment and a nontreated check. Each plot consisted of a single tree. At the Texas location, the experiment also had six trees per treatment and check, but treatments were not randomized; each treatment (or check) consisted of a single large-block in a demonstration-style plot (note that because of lack of randomization among plots the data from the orchard in Texas were not analyzed statistically). The fungi were applied according to Shapiro-Ilan et al. (2008, 2009). B. bassiana was applied as a trunk treatment at all locations, whereas (because of a shortage of material) M. anisopliae trunk-bands were applied at Fort Valley, GA, and Comanche, TX, but not at Byron, GA. For trunk application with B. bassiana, 236.5 ml of BotaniGard ES (5 x 1012 conidia) and 100 ml of SoyScreen oil were mixed with sunflower oil (ConAgra Foods, Irving, CA) to reach a total volume of 1 liter. A CO2-charged backpack sprayer (310.3 kPa, Spray Systems Co., Wheaton, IL) with a cone nozzle (5500-X8 adjustable conejet) was used to apply the mixture to approximately 1.5 m of the bottom part of the trunk. The SoyScreen was added as a UV-protecting adjuvant (Compton and Laszlo 2002). The M. anisopliae fungus bands (approximately 45 x 3 cm) were attached horizontally around the circumference of the trunk by stapling the ends and middle of each; six or seven bands were required to encircle the trunk. Two rings of bands were attached to each tree, one approximately 123 cm above the ground and the other 135 cm above the ground. At Comanche, TX, an aqueous ground application of M. anisopliae also was applied around each tree at a rate of 5 x 1012 conidia per plot (hence, the rate-per-unit area was approximately 6.4 x 1010 conidia per m2); M. anisopliae was mixed with approximately 30.3 liters of water and 0.01% Silwet L-77, (Loveland Industries, Inc., Greeley, CO) and applied via watering can to a radius of 5 m around each trunk. Applications were on 15 August 2006 in Georgia and 24 August in Texas. Efficacy of the treatments was estimated for naturally emerging pecan weevils. Adult pecan weevils were collected in Circle traps attached to pecan trunks (Mulder et al. 2003). This is a passive trap that captures insects crawling up the trunk. The traps were made of 1.5-mm wire mesh with an open area approximately 61 cm wide facing toward the soil (to collect ascending weevils) and tapering up to a removable top. Traps were placed on the trunk so the bottom of the trap was approximately 100 cm above the soil surface.