Effect of Monitoring Technique in Determining the Presence of Fungus Gnat, Bradysia spp. (Diptera: Sciaridae), Larvae in Growing Medium 1

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1 Effect of Monitoring Technique in Determining the Presence of Fungus Gnat, Bradysia spp. (Diptera: Sciaridae), Larvae in Growing Medium 1 Ana R. Cabrera, Raymond A. Cloyd, and Edmond R. Zaborski 2 Department of Natural Resources and Environmental Sciences, University of Illinois, Champaign, Illinois J. Agric. Urban Entomol. 20(1): (January 2003) ABSTRACT Monitoring tools for detecting the presence of fungus gnats (Bradysia spp.) are a critical part of effective IPM programs for these pests in greenhouse production systems. Research has demonstrated the effectiveness of yellow sticky cards for monitoring adult fungus gnats; similarly, potato disks placed on the surface of the growing media in pots have been shown to be effective monitoring tools for the damaging larval stages. Recently, carrot disks have been suggested as an alternative for monitoring fungus gnat larvae, but this practice has not been verified in experiments. Similarly, extension recommendations for using the potato disk method suggest monitoring durations (the time that potato disks are left on the soil surface before examination) varying from 24 to 48 h, but there are no experimental data of the effect of duration on efficiency. The objective of this study was to compare the sampling efficiency of potato and carrot disks and to determine the effect of monitoring duration on efficiency. In a laboratory experiment with 20 third-instar larvae added to peat-based growing media in 250-ml containers without plants, potato and carrot disks did not differ in efficiency, but a greater proportion of larvae were recovered after 48 h (59%) than after 24 h (34%). In a greenhouse experiment with 20 third-instar larvae added to peat-based growing media in 155-mm standard pots with Lantana camara plugs, only 4% of added larvae were recovered, with no difference in efficiency between either potato and carrot disks or 24- and 48-h monitoring durations. When this experiment was repeated using 50 third-instar larvae, the proportion of larvae recovered increased from 16.6% at 24 h to 30.5% at 48 h. With a monitoring duration of 48 h, potato disks recovered significantly more larvae (38.2%) than carrot disks (22.8%). These results suggest that potato disks are more efficient than carrot disks as a monitoring device to detect fungus gnat larvae and also that a monitoring duration of 48 h was more efficient in recovering larvae than 24 h. KEY WORDS disk fungus gnat, Bradysia spp., monitoring, potato disk, carrot Fungus gnats, Bradysia spp., are important insect pests in commercial greenhouse production systems, particularly during propagation and plug production and prior to plants developing well-established root systems (Lindquist 1997, Cloyd 2000). Although both adults and larvae can be a problem, the larvae are 1 Accepted for publication 11 August Illinois Natural History Survey, Champaign, Illinois

2 42 J. Agric. Urban Entomol. Vol. 20, No. 1 (2003) primarily responsible for plant injury. Fungus gnat larvae cause direct plant injury by feeding on plant roots, thus reducing their ability to take up water and nutrients (Jarvis et al. 1993). The larval stages are also capable of disseminating and transmitting diseases (Gardiner et al. 1990, James et al. 1995). In addition, larval feeding causes indirect damage by creating wounds that allow entry of soil-borne plant pathogens such as Pythium spp. (Jarvis et al. 1993, Gardiner et al. 1990). Monitoring devices used to determine the presence or numbers of fungus gnats include the use of yellow sticky cards for adults, and newly cut slices (disks) of potato placed firmly in the growing medium of potted plants for larvae (Lindquist 1997). The potato disk is nondestructive and provides quantitative information in a relatively short period of time (Harris et al. 1995). Alternatively, raw carrot disks have been suggested on some extension web pages (Osborne & Fooshee 2000) as an alternative for monitoring larvae, but the efficacy of carrot disks, particularly in comparison to potato disks, has not been experimentally verified. The suggested monitoring duration (the time that potato or carrot disks are left on the soil surface before examination) for fungus gnat larvae ranges from 24hto 48 h (Cloyd 2000). However, there are no experimental data to demonstrate the effect of monitoring duration on the efficiency of this method. This information would help greenhouse producers use the disk method most effectively for monitoring fungus gnat larval populations. The objective of this study was to compare the efficiency of carrot and potato disks for monitoring larval fungus gnat populations and determine how duration of monitoring influences the efficiency with which fungus gnat larvae are recovered. Materials and Methods Fungus gnats rearing and infestation procedure. Fungus gnats, Bradysia sp., obtained from contaminated pots in a commercial greenhouse in Perryville, Missouri, in 2000 were used to establish a laboratory colony. Colonies were kept in 8-l plastic containers with tight-fitting lids; openings (approximately 15 cm square) were cut in the lids and then covered with fine mesh screen for ventilation. Containers were half filled with Universal Mix (Strong-lite Horticultural Products, Pine Bluff, Arkansas) that was previously heated in a microwave oven at full power (850 W output) for 10 min and enriched with shredded peeled potato (about 1.6 kg). Peeled potatoes were soaked in hot water for 30 min before shredding in a food processor to soften the pulp. Enough water was added to thoroughly moisten the media without having water accumulate at the bottom of the container. Oatmeal (5 ml) was sprinkled over the soil and allowed to mold as an extra food source. Containers were inoculated with several dozen adult fungus gnats collected from previously established culture containers with an aspirator. Each container was kept for only two generations to avoid the build up of high populations of contaminating astigmatid mites. Colonies were maintained under ambient room conditions (21 23 C; approximately 10:14 [L:D] h). To obtain uniformly aged fungus gnat larvae for use in experiments, eggs were collected by placing a 9-cm diameter Petri dish containing moist woodchips into a colony container in which adults had recently emerged. After 24 h, the Petri dish was removed, covered, and held under ambient laboratory conditions. After egg

3 CABRERA et al.: Use of Monitoring Tools for Fungus Gnats 43 hatch (approximately 5 d), the larvae were transferred to plastic 250-ml containers (Fabri-Kal Corp., Kalamazoo, Michigan) half filled with moist Sunshine LC-1 Mix (Sun Gro Horticulture Inc., Bellevue, Washington) and shredded potato. When the larvae reached the third instar ( 7 8 d), they were used to inoculate the experimental units. A small amount ( 10 g) of growing medium containing larvae was placed into a Petri dish with deionized water and, under a dissecting microscope, appropriate numbers of third instars for each experimental unit were counted out and pipetted into 20-ml glass vials. Larvae were rinsed out of vials into experimental units using a wash bottle and deionized water. Laboratory experiment. This trial was set-up under laboratory conditions with the temperature maintained between C. The trial was arranged in a 2 2 factorial complete randomized design with monitoring device (potato vs. carrot disk) and time interval (24 h vs. 48 h) as the treatments measured for effect on the number of fungus gnat larvae recovered. Each treatment combination was replicated 10 times: potato disk at 24 h (P24), potato disk at 48 h (P48), carrot disk at 24 h (C24), and carrot disk at 48 h (C48). Plastic containers (250 ml; Fabri-Kal Corp., Kalamazoo, Michigan) were filled with Sunshine LC-1 Mix. The medium was moistened with tap water to the point that when a handful of moist media was squeezed, approximately 20 drops of water ran off (water content of 4 g g 1 dry media, as determined by oven drying at 60 C). Twenty third-instars were added to each container and allowed to disperse undisturbed within the growing medium for 24 h. After 24 h, a 2.5 cm 2.5 cm (diameter height) freshly cut disk of potato or carrot was placed into the center of each container. The disks were firmly inserted approximately 1 cm into the growing medium. After 24 h or 48 h, depending on the designated treatments, the disks were gently removed from the growing medium. Larvae that were present in or on the potato or carrot disk, as well as any larvae that were present on the growing medium surface beneath the disk, were counted and added, to obtain a total number of fungus gnat larvae for each experimental unit. Greenhouse experiment 1. This trial was set-up in a closed greenhouse section (7.22 m 6.22 m) with the temperature maintained between 18 and 21 C. The trial was arranged as a completely randomized design with 10 replicate pots for each treatment. Lantana camara L. plugs (2.5 cm 2.5 cm) obtained from a commercial greenhouse facility (H. M. Buckley and Sons, Inc., Springfield, Illinois), were planted into 155 mm standard pots (Kord Products, Toronto, Canada) with Sunshine LC-1 Mix. Treatments were randomly assigned to the pots: potato disk at 24 h (P24), potato disk at 48 h (P48), carrot disk at 24 h (C24), and carrot carrol at 48 h (C48). In addition, two control treatments were set-up. The first control consisted of no plant, infested with larvae, and a potato disk left for 48 h. The second control consisted of a plant, no artificial infestation of larvae, and a potato disk left for 48 h. Control one was included to determine the effect of the plant on monitoring efficiency, while the second control was included to check for any larval contamination in the plant growing medium. Twenty third-instars were added to each pot. Larvae were allowed to disperse undisturbed within the growing medium for 24 h and then a 2.5 cm 2.5 cm (diameter height) freshly cut potato or carrot disk was placed approximately 2.5 cm from the plant stem, with approximately 1 cm inserted into the growing medium. After 24 or 48 h, depending on the treatment, the potato or carrot disks

4 44 J. Agric. Urban Entomol. Vol. 20, No. 1 (2003) were gently lifted from the growing medium. Larvae that were present in or on the potato or carrot disk, as well as any larvae that were present on the growing medium surface beneath the disks, were counted and added, to determine the total number of fungus gnat larvae for each replicate. Greenhouse experiment 2. The first greenhouse experiment was repeated using a density of 50 third-instars per pot; replications were blocked through time. Data analysis. All data were analyzed using the Statistical Analysis System version 8.2. Data were tested for normality with the PROC UNIVARIATE procedure, and an analysis of variance (ANOVA) was conducted for the laboratory and greenhouse experiments using the PROC MIXED procedure (SAS Institute 1999). Because the comparisons of interest (5) in the greenhouse experiments were nonorthogonal and exceeded the available degrees of freedom, a Bonferroni adjustment was made to maintain the experimentwise Type I error rate at The contrasts of interest were: main effect of monitoring device (potato vs. carrot), main effect of time (24 h vs 48 h), the effect of the presence of a plant (P48 vs control-1), the effect of monitoring device at 24 h (P24 vs C24), and the effect of monitoring device at 48 h (P48 vs. C48). Results and Discussion Laboratory experiment. No significant interaction between the two monitoring devices (potato or carrot) and time (24 h or 48 h) was detected (P 0.4). In addition, no significant difference between the use of potato or carrot disks (P 0.2) was found. However, there was a significant difference (P < ) in the time intervals (24 h vs. 48 h) that the potato and carrot disks were allowed to remain on the growing medium. Leaving potato and carrot disks for 24 h in the growing medium resulted in recovering an average of 6.6 and 7.0 fungus gnat larvae, respectively (Table 1); this represents a 33 to 35% efficiency. However, when the potato and carrot disks were left for 48 h in the growing medium this resulted in recovering an average of 10.9 and 12.5 fungus gnat larvae, respectively (Table 1), a 54.5 and 62.5% efficiency. Leaving the disks for longer periods of time may have allowed the larvae to detect odors emitted from the disks, giving the larvae time to locate them. In addition, the size of the container may have restricted larval distribution such that more larvae were located in closer proximity to the potato and carrot disks. Greenhouse experiment 1. No significant effect of the model (P 0.08) was found. These results may be explained by the low number of larvae (< 1 larvae per 155 mm standard pot) recovered in the trial, which made it difficult to detect any differences in the treatments (Table 2). The container size may have influenced the results, compared to the laboratory study, by providing more space for the larvae to disperse out into the soil profile. As a result, fewer fungus gnat larvae may have been located close enough to the potato or carrot disks to be attracted to them. Three fungus gnat larvae were detected in one pot in the second control (plant, no fungus gnats added) treatment, indicating contamination of the Lantana plugs used in the experiment. Greenhouse experiment 2. A significant difference (P ) was found when comparing potato and carrot disks in monitoring for fungus gnat larvae with a density of 50 fungus gnat larvae per pot. On average, 5.7 more larvae were

5 CABRERA et al.: Use of Monitoring Tools for Fungus Gnats 45 Table 1. Number of fungus gnat, Bradysia sp., larvae recovered from potato and carrot disks after 24 and 48 h from 250-ml containers of peat-based growing medium infected with 20 third-instar fungus gnat larvae. Treatment 24 h 48 h Potato 6.60 ± 0.65 a ± 0.85 Carrot 7.00 ± ± 0.85 a Data are means ± 1SE, n 10. recovered from the growing medium when using potato disks compared to carrot disks. This represents an 11.5% increase in efficiency of the monitoring device. In addition, when comparing the main effect of time (24 h or 48 h) there was an average increase (P ) of 7.0 in the number of larvae recovered when the potato or carrot disks were left for 48 h compared to 24 h. This represents a 14% increase in efficiency of the monitoring technique (Table 3). After 24 h, no significant difference between potato and carrot disks was found (P 0.1), but there was a significant difference after 48 h (P ). The average number of larvae recovered after 48 h when using potato disks was 19.1, a 38.2% efficiency (Table 3). The recovery of larvae using potato disks for 48 h was not influenced by the presence of a plant (P 0.7). This may be explained by the fact that when a plant is present, fungus gnat larvae may inhabit areas near the root system, whereas in the absence of a plant, the larvae may disperse deeper into the growing medium profile in search of a food source. However, these results may differ when using another size pot, a different size plant, or another plant species. In addition, although most fungus gnats are located in the upper 1 2 inches of the growing medium (Rutherford et al. 1985), some larvae may be found throughout the grow- Table 2. Number of fungus gnat, Bradysia sp., larvae recovered from pots of peat-based growing media in the greenhouse using potato and carrot disks; influence of monitoring duration and presence of a Lantana plant (Lantana camara). Plant Larvae added Duration (h) Number of larvae recovered Potato disk Carrot disk Lantana ± 0.55 a 0.00 ± 0.00 Lantana ± ± 0.41 Lantana ± 0.30 n.d. b None ± 0.43 n.d. a Data are means ± 1 SE, n 10. b n.d.: not determined.

6 46 J. Agric. Urban Entomol. Vol. 20, No. 1 (2003) Table 3. Number of fungus gnat, Bradysia sp., larvae recovered from pots of peat-based growing media in the greenhouse using potato and carrot disks; influence of monitoring duration and presence of a Lantana plant (Lantana camara). Plant Larvae added Duration (h) Number of larvae recovered Potato disk Carrot disk Lantana ± 1.6 a 6.4 ± 1.0 Lantana ± ± 1.7 Lantana ± 0.0 n.d. b None ± 2.4 n.d. a Data are means ± 1SE, n 10. b n.d.: not determined ing medium profile (Zaborski & Cloyd, unpublished data). This may influence the accuracy of the monitoring technique. No fungus gnat larvae were recovered from the second control treatment (plant, no fungus gnat larvae added), indicating no contamination of the plant plugs used in this experiment. The use of potato disks as a monitoring device to detect the presence and the numbers of fungus gnats in pots has been the primary recommendation to greenhouse producers (Lindquist 1997, Cloyd 2000). The results of this experiment suggest that the use of carrot disks is not as efficient as potato disks. A monitoring duration of 48 h with potato disks gave an almost 20% increase in efficiency compared to 24 h. Thus, a monitoring duration of 48 h is recommended because the detection of larvae, especially at low population densities, is more likely. It was observed that the recovery efficiency of the monitoring technique differed between greenhouse experiments 1 and 2. The percentage recovery was higher at a high density (50 larvae pot 1 ) than at a low density (20 larvae pot 1 ). This variation could have been caused by factors such as the environment, larval density, larval instar, or competition of the larvae for root area. This may indicate that the use of potato disks would be more reliable for detecting the presence of fungus gnat larvae in the soil-less medium than to estimate absolute numbers of larvae. Acknowledgments We thank Amy Dickinson for maintenance of fungus gnat colonies and Erick Caamaño, Monica Garces, Janeth Moncada, and Carlos Pavon for technical assistance. This research was supported by a grant from the Illinois Department of Agriculture (Sustainable Agriculture Grant Program, no. SA00-11) to RAC and ERZ. References Cited Cloyd, R. A Fungus gnat and shore fly management strategies: Panel discussion, pp In Proceedings for the 16 th Conference on Insect and Disease Management on Ornamentals. Society of American Florists, Alexandria, Virginia. 227 pp.

7 CABRERA et al.: Use of Monitoring Tools for Fungus Gnats 47 Gardiner, R. B., W. R. Jarvis & J. L. Shipp Ingestion of Pythium spp by larvae of the fungus gnat Bradysia impatiens (Diptera: Sciaridae). Ann. Appl. Biol. 116: Harris, M. A., R. D. Oetting & W. A. Gardner Use of entomopathogenic nematodes and a new monitoring technique to control fungus gnats, Bradysia coprophila (Diptera: Sciaridae), in floriculture. Biol. Control 5: James, R. L., R. K. Dumroese & D. L. Wenny Botrytis cinerea carried by adult fungus gnats (Diptera: Sciaridae) in container nurseries. Tree Planters Notes 46: Jarvis, W. R., J. L. Shipp & R. B. Gardiner Transmission of Pythium aphanidermatum to greenhouse cucumber by the fungus gnat Bradysia impatiens (Diptera: Sciaridae). Ann. Appl. Biol. 122: Lindquist, R. K. July Integrated pest management of poinsettia pests: Fungus gnats. Ohio Florists Assoc. Bull. 813: 5 8. Osborne, L. S. & W. C. Fooshee Fungus Gnats. University of Florida, Mid-Florida Research and Education Center, Apopka, Florida From the Wide World Web: Accessed: December 13, Rutherford, T. A., D. B. Trotter & J. M. Webster Monitoring fungus gnats (Diptera: Sciaridae) in cucumber greenhouses. Can. Entomol. 117: SAS Institute SAS OnlineDoc, Version 8.02 SAS Institute, Cary, North Carolina.