W. Michael Hood, Paul M. Horton, and John W. McCreadie 2. Department of Entomology, Clemson University, Clemson, South Carolina

Transcription

1 Field Evaluation of the Red Imported Fire Ant (Hymenoptera: Formicidae) for the Control of Wax Moths (Lepidoptera: Pyralidae) in Stored Honey Bee Comb 1 W. Michael Hood, Paul M. Horton, and John W. McCreadie 2 Department of Entomology, Clemson University, Clemson, South Carolina J. Agric. Urban Entomol. 20(2): (April 2003) ABSTRACT The red imported fire ant, Solenopsis invicta Buren, was studied as a potential biological control agent of wax moths (the greater wax moth, Galleria mellonella L. and the lesser wax moth, Achroia grisella F.) in stored beehive supers of drawn comb. This 3-yr study evaluated three methods of super storage arrangement on experimental sites having red imported fire ant colony densities ranging from 107 to 393 mounds per ha. Wax moth damage to comb was significantly less on equipment stored on wooden pallets exposed to increased light, greater ventilation, and sites having 214 red imported fire ant mounds per ha when compared with equipment stored similarly with ants excluded. Predation against wax moths was found to increase with an increase in red imported fire ant colony density. The red imported fire ant is a promising biological control agent of wax moths, but was found to be consistently effective only at excessive colony densities when equipment was stored in a manner which promoted increased light and ventilation conditions. Beekeepers may accept marginal wax moth control in stored comb placed in areas of less red imported fire ant colony density when supers are stacked in a manner that promotes light and ventilation. A vertical stack of supers of comb in a normal beehive arrangement that allowed poor ventilation and light conditions resulted in excessive wax moth damage even at the highest fire ant colony density (331 mounds per ha). KEY WORDS Hymenoptera, Formicidae, Solenopsis invicta, red imported fire ants, Lepidoptera, Pyralidae, Galleria mellonella, wax moths, biological control, pest management The greater wax moth, Galleria mellonella L. (Lepidoptera: Pyralidae) is a serious honey bee pest in the warm regions of the world (Calvert 1982). The greater wax moth is an important pest of several honey bee species, including Apis mellifera L. (Hymenoptera: Apidae), A. cerana F., A. dorsata F., and A. florea F. (Southwick 1987). Nests of non-apis bee species affected by the greater wax moth include meliponids (Nogueiro-Neto 1953) and bumble bees (Oertel 1963). The lesser wax moth (Achroia grisella F.) is less known for its damage to beekeeping equipment but is often found feeding in the same wax comb as the greater wax moth. Adult wax moths and larvae are capable of transferring serious bee diseases, such as foulbrood (Charriere & Imdorf 1999). 1 Accepted for publication 27 February Department of Biological Sciences, University of South Alabama, Mobile, Alabama,

2 94 J. Agric. Urban Entomol. Vol. 20, No. 2 (2003) The greater wax moth can completely destroy unprotected comb. Dark, aged comb containing many bee larval cocoons are most vulnerable (Charriere & Imdorf 1999). Significant investment in labor and capital are required annually by southern United States beekeepers to replace combs destroyed by this pest (Williams 1976). Wax moth infestation of comb honey results in contamination of a delicate product that is unmarketable (Burges 1978). Estimated losses from the greater wax moth were $3 million in 1973 and $4 million in 1976 to 114 southern United States commercial beekeeping operations. Estimated losses from the greater wax moth were $1.50 per colony in Texas and $3 per colony in Florida. National annual losses to this pest have been estimated at $5 million or more (Williams 1997). In a 1991 survey of beekeepers in Georgia, respondents listed the greater wax moth as the most serious honey bee problem in their apiary operations (Delaplane 1991). Several chemical and nonchemical methods have been developed to control wax moths in stored comb. Freezing, heating, and the use of carbon dioxide as a fumigant to protect stored equipment are examples of nonchemical control, but their use is often unrealistic for the commercial beekeeper. Paradichlorobenzene is a fumigant currently registered in the United States for wax moth control in stored equipment, but the safety of this product is questioned (Wallner 1992, Charriere & Imdorf 1999). Other chemical fumigants such as acetic acid, calcium cyanide, ethylene dibromide, methyl bromide, and phosphine have been successfully used to control wax moths in stored equipment (Burges 1978, Cantwell 1980, Bailey 1981, Shimanuki 1981, Goodman et al. 1990). Of these chemicals, only phosphine (Phostoxin ) is approved in the United States as a restricted-use pesticide to control wax moths in empty comb and comb honey to be used as bee food (Williams 1997). The microbial product Certan, which contains B. thuringiensis, provides excellent wax moth control on stored equipment (Cantwell & Shieh 1981), but it is no longer commercially available in the United States. A safe, low-cost method of wax moth control is needed, especially for commercial beekeepers operating in the tropics and subtropics. Trichogramma sp. (Hymenoptera: Trichogrammatidae) have been reported to parasitize wax moth eggs successfully, but inspection of newly stored comb for moth larvae is necessary (Bollhalder 1999). The red imported fire ant (RIFA) Solenopsis invicta Buren (Hymenoptera: Formicidae), an omnivorous and highly effective insect predator (Arant et al. 1958, Hays & Hays 1959, Horton et al. 1975, Vinson 1997), preys on immature stages of the greater wax moth (Williams 1997). Fire ants establish a network of underground tunnels that radiate from the mound and have been reported to cover a foraging range of 53 to 94 m 2 (Markin et al. 1975). The RIFA now infests over 106 million ha (275 million ac) of land, primarily in nine southeastern states of the United States and Puerto Rico, with small infestations in Oklahoma and Tennessee (Vinson 1997). The overlapping generations of wax moths and the RIFA offer the potential for beekeepers to take advantage of this natural predator-prey relationship. Typically, beekeepers in the southern region of the United States extract their honey in summer and store their supers of empty-drawn comb at this time till the following spring. Summer and fall are the most vulnerable time periods of the year for wax moth activity in stored comb. In this article, we present the results of field studies on the effectiveness of the red imported fire ant in controlling wax moths in stored supers of comb under variable light and ventilation conditions.

3 HOOD ET AL.: Biological Control of Wax Moths 95 Materials and Methods Experimental sites were selected in four South Carolina counties, which had variable numbers of RIFA colonies per ha. Experimental sites were located in the Piedmont and Sandhill regions of the state and were pastureland or undisturbed field sites that had full sunlight and no overhead vegetation, which substantially increases RIFA foraging activities (Porter & Tschinkel 1987). No attempt to control fire ants on the four experimental sites had been conducted since RIFA first appeared in the Sandhill and Piedmont regions of South Carolina about 35 years previously. The number of RIFA colonies per ha was measured at each site during initial test plot establishment and at the end of the 90-d tests (Table 1) by counting the active RIFA mounds within a circular plot having an 18.3 m (60 ft) radius and multiplying this number by (ex. 10 active RIFA mounds within the 18.3 m radius of circular plot center mounds per ha). A RIFA colony was considered active if ants exited the mound after disturbance of the mound surface with a plastic probe. Experimental sites were visited on a 30-d interval to monitor RIFA site activity, inspect comb for RIFA damage, and measure wax moth damage. Wax moth damage to comb was identified by the presence of a mass of webbing and debris whereas RIFA damage was identified by the presence of a powdery residue with no webbing. Supers of drawn comb, where honey had been extracted and exposed to allow foraging bees to remove honey residues over a 2-wk period, were placed in a chest-type freezer for 3 h at 12 C (10 F) to kill all life stages of wax moths (Shimanuki & Knox 1997). Each frame of comb was removed from supers and inspected for signs of wax moth activity. Only frames showing no signs of past wax moth activity were used in tests. A mixture of aged, dark comb frames and newly drawn comb frames were used in tests. Nine frames were randomly se- Table 1. Pre- and post-test active red imported fire ant (RIFA) colony mounds per ha (RIFA mounds per ac in parentheses) during the 90-d treatment periods for three experimental sites in South Carolina, Location Precount Postcount Test 1 Greenville County 173 (69) 106 (43) Laurens County 192 (78) 215 (87) Saluda County 331 (134) 183 (74) Test 2 Greenville County 148 (60) 84 (34) Laurens County 128 (52) 106 (43) Saluda County 393 (159) 264 (107) Test 3 Laurens County 107 (43) 64 (26) Richland County 214 (87) 222 (90) Saluda County 299 (121) 264 (107)

4 96 J. Agric. Urban Entomol. Vol. 20, No. 2 (2003) lected and placed in each test super to promote equal attraction of wax moths in the field. Each super of nine frames was considered a replication in experiments 2 and 3. Greater wax moth colonies were reared in 10-frame-deep Langstroth hive bodies ( cm) with frames of aged comb without bees present. Each test super of drawn comb was infested with greater wax moth by placement of three late instar larvae on frames at field placement to insure wax moth presence at each test site although exposed beeswax is highly attractive to wax moth adults in the southern United States. Experiment 1 (year 1). In the first week of August 1995, we established experimental sites in Greenville, Laurens, and Saluda Counties where RIFA mound activity was measured (Table 1). Four treatments of stacked 10-framedeep Langstroth hive bodies with telescoping hive tops and bottom boards were placed on each experimental site. Hive bodies were stacked in a normal beehive manner (Fig. 1) that allowed only minimum light and ventilation through the bottom entrance, simulating a dead colony. Variable numbers of stacked hive bodies were used at each site to test different degrees of wax moth comb destruction from the bottom to the top hive body. The treatments were: 1) four hive bodies stacked on two concrete blocks ( cm), 2) two hive bodies stacked on two concrete blocks, 3) one hive body placed on two concrete blocks, and 4) one hive body placed on a wooden table (height-48 cm, width-38 cm, length-65 cm) with each of the four legs resting in a number 10 round vegetable can (height-18 cm, diameter-16 cm), half filled with used motor oil to exclude RIFA. Experiment 2 (year 2). In the last week of July 1996 we established experimental sites in Greenville, Laurens, and Saluda Counties where RIFA mound Fig. 1. Stacking arrangement for supers of comb frames in experiments 1, 2, and 3 that evaluated red imported fire ants for controlling wax moths in South Carolina.

5 HOOD ET AL.: Biological Control of Wax Moths 97 activity was measured (Table 1). Five treatments of positioned four shallow supers ( cm) with nine frames each on end in a horizontal super stack orientation rather than the normal vertical stack beehive orientation were placed on each experimental site. A nylon hive strap with ratchet was used to secure the four supers together for each treatment (Fig. 1). Supers were placed together with frame top bars facing inward to prevent frames from falling out of the supers. This super arrangement allowed increased light and ventilation compared with experiment 1. A piece of plywood ( cm) was placed over the supers to protect equipment from rain and direct sunlight. Treatments were: 1) supers stored on a wooden pallet with a wood bound wire queen excluder attached on each end to exclude rodents and other small pests, but allow light, ventilation, and access by RIFA (L/V/I); 2) supers stored on a wooden pallet with a solid hive cover ( cm) secured on each end to minimize light and ventilation, but allow RIFA access (IFA); 3) supers stored on a table with legs resting in number 10 vegetable cans half filled with used motor oil to exclude RIFA and a piece of plexiglas ( cm) secured on each end of the supers to minimize ventilation, but to allow entry of light (L); 4) supers stored on a table with legs resting in number 10 vegetable cans half filled with used motor oil to exclude RIFA and a wood bound wire queen excluder secured to each end of supers to exclude rodents and other small pests (L/V); (5) supers stored on a table with legs resting in number 10 round vegetable cans half filled with used motor oil to exclude RIFA and a solid hive cover secured to each end to minimize light and ventilation (control). Experiment 3 (year 3). In the second week of August 1997 we established experimental sites in Laurens, Richland, and Saluda Counties where RIFA mound activity was measured (Table 1). Three treatments of five supers stacked vertically with frames of adjacent supers alternated in a crisscrossed fashion were placed on each experimental site (Fig. 1). A piece of plywood ( cm) was placed over the top super to protect equipment from rain and direct sunlight. The bottom super was exposed without a bottom board. Supers were positionedto allow light and ventilation to enter the comb from top to bottom super. The treatments were 1) supers stacked on a wooden pallet to allow light and ventilation and to allow access by RIFA (L/V/I), 2) supers stacked on a wooden pallet with a white cloth sheet draped over the equipment to reduce light and ventilation, but to allow access by RIFA (IFA), and 3) supers placed on a table with legs resting in number 10 round vegetable cans, half filled with used motor oil to exclude RIFA (L/V). Data collection. Wax moth damage was measured 90 d after the initiation of each experiment. Damage estimates were made by placing a piece of plexiglas ( cm) with a scribed 1-cm grid over each side of test frames. Damage was estimated by counting the total number of cm 2 of comb destroyed by wax moths on each side of frames. Damage estimates were recorded without attempting to differentiate damage resulting from the two species, the greater wax moth or the lesser wax moth. Data analyses. Initial examination of the damage data showed that sample variances far exceeded sample means and thus a log (Y + 1) transformation (where Y damage in cm 2 ) was necessary (Ott 1993). Both two-way and one-way analysis of variance (ANOVA) were performed on the log-transformed estimates of wax moth damage. Main effects were treatment (five levels) and experimental

6 98 J. Agric. Urban Entomol. Vol. 20, No. 2 (2003) site (three levels). For significant ANOVA (P < 0.05), Dunnett s test was used to determine if treatment means were significantly different from the control means (experiment 2). Because we were interested in whether treatments promoted or reduced wax moth damage, a two-tailed test was used (Zar 1984). Tukey s multiple comparison for differences among treatment means was used for experiment 3. Results Experiment 1. There was no significant (P > 0.05) difference in wax moth damage among treatments regardless of RIFA colony density when hive bodies were stored in a normal beehive arrangement. Comb was severely damaged (>90%) from top to bottom hive body even at the highest RIFA density experimental site. Experiment 2. Both treatment and experimental site had a highly significant (P < 0.001) affect on wax moth damage in supers (Table 2). However, given the highly significant (P < 0.001) interaction effect (treatment x experimental site), multiple comparisons among treatment levels for each factor (treatment, experimental site) were not considered appropriate (Zar 1984). Thus, results were re-analyzed for each experimental site using a one-way ANOVA (main effect treatments) with significant ANOVA further analyzed using Dunnett s test (Table 2). For Saluda County, the L/V treatment had significantly greater (P 0.029) and the L/V/I treatment had significantly less (P 0.039) wax moth damage than the control. For this experimental site, our data indicate that a combination of RIFA, ventilation, and light resulted in substantial wax moth control. Light and ventilation only, without the presence of RIFA, resulted in increased wax moth damage. In Laurens County, both L and L/V/I treatments had significantly higher (P and P , respectively) damage than the control (Table 3). Treatment had no significant affect on moth damage in Greenville County (Table 3). Experiment 3. Both treatment and experimental site had a highly significant (P < 0.001) affect on wax moth damage in supers (Table 4). However, as with experiment 2, there was a highly significant (P < 0.001) interaction effect (treatment x experimental site). Thus, results were analyzed for each experimental site using a one-way ANOVA (main effect treatments) with significant ANOVA and differences among treatments further analyzed using Tukey s test (Table 5). For Table 2. Two-way analysis of variance on log-transformed estimates of wax moth damage with five treatments on three experimental sites having varying numbers of red imported fire ant colonies in South Carolina, 1996 (experiment 2). Source of variation DF MS F P Treatments <0.001 Site <0.001 Treatment site <0.001 Error Total 59

7 HOOD ET AL.: Biological Control of Wax Moths 99 Table 3. One-way analysis of variance (ANOVA) and Dunnett s multiple comparison analysis of mean wax moth damage (cm 2 ) with differing combinations of light, ventilation, and red imported fire ant (RIFA) colony densities on three experimental sites in South Carolina, 1996 (experiment 2). Dunnett s test was performed only on those experimental sites in which the ANOVA was significant (P < 0.05). Treatments a Log data Damage (cm 2 ) b Raw data Saluda County <0.001 Control IFA L L/V 1.11** L/V/I 0.21* 1.43 Laurens County <0.001 Control IFA L 1.92** L/V L/V/I 1.47** Greenville County c 2.98 <0.054 a Treatment abbreviations: IFA, RIFA only; L, light only; L/V, light and ventilation only; L/V/I, light, ventilation and RIFA; Control, no light or ventilation and RIFA excluded. b All analyses were performed on the log-transformed data; raw data are given for comparison purposes. For each experimental site, * indicates a mean < the control, and ** indicates a mean > than the control. c The ANOVA was not significant for this experimental site, Dunnett s test was not performed. F P Saluda County, the L/V, IFA and L/V/I treatments were all significantly different from each other with L/V showing the greatest damage and IFA treatment having the least (Table 5). In Richland County, the L/V/I treatment had significantly less damage than either the L/V or IFA treatments (Table 5). Treatment had no significant affect on moth damage in Laurens County (Table 5). Discussion Although the RIFA is considered by humans to be a major pest, results from our investigations indicate that this insect can be used at high colony density levels to provide excellent wax moth control in stored supers of comb, if measures are taken to promote light and ventilation. These experiments were initiated in late July or early August, which is the time of year when many southern United States beekeepers extract honey and store empty supers of comb frames. This is a time of year when wax moths are abundant and the comb is vulnerable to wax moth damage. Also, this is the time of year when RIFA worker populations increase during the second half of the year (Tschinkel 1993), and brood food (pro-

8 100 J. Agric. Urban Entomol. Vol. 20, No. 2 (2003) Table 4. Two-way analysis of variance on log transformed estimates of wax moth damage with three treatments on three experimental sites having varying numbers of red imported fire ant colonies in South Carolina, 1997 (experiment 3). Source of variation DF MS F P Treatments <0.001 Site <0.007 Treatment site <0.001 Error Total 44 tein) is necessary for colony growth (Porter & Tschinkel 1987). Many beekeepers protect their stored comb from wax moths by chemical treatment. Beekeepers who live in the RIFA infested areas may benefit greatly by using this safe, economical method of biological control of wax moths. The high RIFA colony density levels required ( 214 colonies per ha in experiment 3 and 393 colonies per ha in experiment 2 may limit the use of this biological control method to rural areas where other human or animal activities are limited Table 5. One-way analysis of variance (ANOVA) and Tukey s multiplecomparison analysis of mean wax moth damage (cm 2 ) with differing combinations of light, ventilation, and red imported fire ant (RIFA) colony densities on three experimental sites in South Carolina in 1997 (experiment 3). Tukey s test was performed only for those experimental sites in which the ANOVA was significant (P < 0.05). Treatments a Log data Damage (cm 2 ) b Raw data Saluda County <0.001 L/V 1.32a IFA 0.13c 0.35 L/V/I 0.66b 3.10 Richland County 8.00 <0.006 L/V 1.45a IFA 1.38a L/V/I 0.55b 2.93 Laurens County c 1.76 <0.214 a Treatment abbreviations: L/V, light and ventilation only (Control); IFA, RIFA only; L/V/I, light, ventilation, and RIFA. b All analyses were performed on the log-transformed data; raw data are given for comparison purposes. For each county, the same letters indicate means not significantly different from each other at a family error rate of P < c The ANOVA was not significant for this county, Tukey s test was not performed. F P

9 HOOD ET AL.: Biological Control of Wax Moths 101 and RIFA are not controlled regularly. Beekeepers may experience increased but acceptable levels of wax moth damage on equipment in areas of reduced RIFA mound density levels (<214 RIFA mounds per ha) if increased light and ventilation conditions are promoted as in experiments 2 and 3. Among the three methods tested for storing supers of comb for wax moth control, the vertically stacked supers that alternated orientation with front forward then side forward (experiment 3) or horizontally stacked supers with frames positioned vertically (experiment 2) provided excellent wax moth control consistently, but only at high RIFA ant mound densities when increased light and ventilation were promoted. The vertical stack (experiment 1) in a normal beehive arrangement that allowed poor ventilation and light conditions resulted in excessive wax moth damage even at the highest RIFA colony density. Therefore, a beekeeper who does not remove dead, extremely weak, or non-active bee colonies from apiaries during the wax moth s active season may realize major comb destruction even at high RIFA colony densities in the warmer regions of the world. Periods of RIFA foraging activity may vary considerably in different areas of the S. invicta range in the southern United States. Shorter foraging activity periods are expected in higher latitudes and distinctly shorter periods in shady habitats (Porter & Tschinkel 1987). Quantity and quality of available food (protein) will likely play a dominant role in RIFA foraging activities, especially during colony brood rearing periods. Therefore, temporary storage of combs that are heavily infested with wax moths near RIFA mounds may result in heavy predation during July-September. RIFA are opportunists (Vinson 1997) and will vary in their foraging activities depending on available food during suitable weather conditions. Reduced RIFA foraging can be expected during extreme temperatures, rainfall, and other weather conditions that may negatively affect this biological control method. Season is an important predictor of RIFA foraging, and reduced foraging activity is expected in late fall (Porter & Tschinkel 1987) during cooler weather when wax moth activity declines, also. We experienced a general, seasonal decrease in number of RIFA colonies during these tests (Table 1), but the remaining active RIFA colonies are expected to have maximum populations in November (Tschinkel et al. 1995), which should provide some wax moth control even in cooler weather. Treatment placement on experimental sites was random and we intentionally positioned treatments at least 2-m from active RIFA mounds to avoid colony disturbance. On some sites, RIFA colonies established mounds adjacent or underneath the equipment during the 90-d field tests. In our tests, all equipment was placed in the field having no wax moth life stages except the three larvae inserted per super at field placement. This low prey density may have delayed the development of RIFA foraging activity patterns in those locations, especially in areas of reduced mound densities (<214 RIFA mounds per ha). We have begun further studies in the use of RIFA attractants placed in supers of comb at field placement to increase ant foraging activities over extended periods to enhance wax moth biological control. Our future research goal is to increase RIFA foraging activity, especially on experimental sites with low mound densities by placement of a RIFA attractant inside supers of stored comb. The presence of pollen and honey in comb may stimulate RIFA feeding which could lead to comb destruction, especially if other natural food is unavailable.

10 102 J. Agric. Urban Entomol. Vol. 20, No. 2 (2003) Therefore, it is important that beekeepers place only dry supers that are free of residual honey in the field for wax moth control as described in these experiments. No RIFA comb damage or comb degeneration due to light and air exposure was observed during these tests. Other ant species (non-rifa) were not observed foraging on the equipment during our periodic visits to the test sites. The beekeeper must be careful when handling equipment that has been stored in RIFA-infested areas. The use of protective gloves is recommended when handling equipment to prevent RIFA stings. Anaphylactic shock is a possible side effect to a RIFA sting for less than 1% of the human population (Vinson 1997). Therefore, we highly discourage the use of this practice to beekeepers that have experienced a severe reaction to RIFA stings. The beekeeper should delay handling and removal of the equipment from a RIFA field-storage site to cold storage till late fall or early winter when low temperatures severely reduce RIFA foraging. Also, the winter season is a time when wax moth activity is minimal. Our results indicate that light and ventilation play an important role when using the RIFA as a biological agent for controlling wax moths in stored supers of comb. Popolizio & Pailhe (1973) reported highly satisfactory wax moth control when frames of comb were stored in wax moth safe storage rooms in Argentina. Comb frames were removed from supers and stored on racks at 3-cm intervals in a shelter with a ceiling only. They made no reference to the presence of RIFA or other insect predators and attributed wax moth control to conditions of light and ventilation. NOTE: We recommend this method of wax moth biological control be used only in areas of the United States that are currently infested with RIFA. The RIFA is a regulated insect pest in the United States and the movement of RIFA or infested associated material or equipment outside the regulated zone is prohibited in many states (Flottum 2002). Acknowledgments The authors thank Larry Grimes (Experimental Statistics Department, Clemson University) for assistance in part of the statistical analyses and Clemson University Sandhill Experiment Station, L.C. Reynolds, Bill Gentry, and Carl Bagwell for the use of their farm sites. Partial funding for this research was provided by the Clemson University Extension IPM Grant program. This is Technical Contribution No of the South Carolina Agricultural Experiment Station. The authors thank John Morse and Al Wheeler (Department of Entomology, Clemson University) for their review and valuable comments on this manuscript. References Cited Arant, F. S., K. L. Hays & D. W. Speake Facts about the imported fire ant. Alabama Highlights Agric. Res. 5: Bailey, L Honey bee pathology. Academic Press, London, 124 pp. Bollhalder, F Trichogramma for wax moth control. Am. Bee J. 139: Burges, H. D Control of wax moths: physical, chemical and biological methods. Bee World 59: Calvert III, P Certan, a bacterial insecticide for control of wax moth (a literature review). Am. Bee J. 122:

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