Invasive and Exotic Species

Chapter 1: Biological Control Agents

Biological Control of Hemlock Woolly Adelgid

Pathogens - Bruce L. Parker, Scott Costa, Margaret Skinner

From: Cheah, C., M. E. Montgomery, S. Salom, B. L. Parker, S. Costa, and M. Skinner, 2004. Biological control of hemlock woolly adelgid. USDA For. Serv. FHTET-2004-04, Reardon, R. and B. Onken (Tech. Coordinators), 22pp

Insect pathogens (entomopathogens) can dramatically reduce insect populations (e.g., declines in gypsy moth populations in North America due to the fungus Entomophaga maimaiga). Most entomopathogens are microorganisms and include fungi, bacteria, viruses, protozoa, rikettsia and microsporidia.

Insect-killing Fungi

Insect-killing fungi differ from most entomopathogens because they can penetrate directly through the body wall of an insect (Fig. 16). Following the fungus-induced death of an insect, large numbers of fungal spores are released from the cadaver to continue the infection cycle throughout the insect population.

Figure 16. Generalized infection cycle of insect-killing fungi.

There is a long history associated with the use of insect-killing fungi for insect pest management. Four notable species, Beauveria bassiana, Metarhizium anisopliae, and Verticillium lecanii (Fig. 17), and Paecilomyces sp., have a nearly ubiquitous, terrestrial, worldwide distribution. Although these fungi are generalists in their host range, a variety of biological, ecological, and behavioral factors serve to limit their effects on non-target species. Also, certain species and some isolates of a given species tend to be virulent to insects within a given order.

Thousands of HWA, many showing signs of fungal infection, were collected from hemlock forests along the eastern seaboard of the United States and from southern China (Reid et al. 2002). From those collected, 79 different insect-killing fungal isolates were recovered, established in pure culture, and identified to species. Pure cultures have been placed in long-term storages at the University of Vermont Entomology Research Laboratory (ERL), Burlington, Vermont, and at the USDA, Agriculture Research Service (ARS), Ithaca, New York.

Extensive laboratory studies have been done to verify that these and other isolates from the ERL collection do indeed infect HWA (Reid 2003). Several of the more virulent isolates were selected for further testing. The selected isolates readily germinated and infected HWA at temperatures normally found throughout HWA’s range and where hemlocks commonly grow. Field tests have been conducted to determine optimal formulations and spray delivery systems, rates, and timings (e.g., spring vs. fall). The effects of fungi on S. tsugae, a non-target predator of HWA, and the development of technology for mass production of fungi for field-testing also are being studied. Although research is on-going, considerable progress has been made and results indicate fungi have good potential as a biological control agent for managing HWA.

Field Efficacy of Insect-Killing Fungi

Between spring 2001 and fall 2003, various field trials were conducted at Mount Tom Reservation, Holyoke, Massachusetts, to select the most virulent fungal isolates and determine the best timing and concentration for fungus applications. Fungi were applied to HWA-infested branches using hand-held and pressurized, and ULV sprayers (five to six branches per treatment). Both single and multiple applications of conidia and blastospores were tested at concentrations ranging from 5 x 107 to 2 x 108 spores per milliliter. Non-treated and formulation blank treated controls were used. Horticultural oil (1.0 percent) was used as a positive control. Survival and density of HWA were examined 3 to 4 weeks after application.

A reduction in HWA populations was selected for gauging fungal efficacy, because it is more reliable than observed mortality, and it best reflects our goal of suppressing HWA populations. During 2002 and 2003, HWA populations were significantly reduced by a single fall application of ARSEF 6010 (V. lecanii) at 1 x 108 spores per milliliter (Fig. 18, page 15). The presence of fungal outgrowth on an HWA cadaver killed by V. lecanii suggests the fungus could re-infect additional insects (Fig. 19, page 15). A second isolate, CA 603 (B. bassiana), had significant activity only during 2002, and was also effective when applied using a ULV sprayer (data not presented).

Horticultural oil also caused substantial population reductions, demonstrating the utility of the population-based approach for evaluating fungal efficacy. Similar applications of either fungi or oil during spring HWA population build-up were not as effective. The reduced effectiveness might be due to the fact that, in spring, HWA are covered with a waxy wool that might prevent the spray from making direct contact with the insect. The wool is absent during late summer and fall when HWA are aestivating as first instar sistens. Targeting HWA during late summer and fall also provides a wider window of opportunity for applying fungi.

Figure 17a. Beauveria bassiana Spores Photo by Svetlana Y. Gouli University of Vermont

Figure 17b. Metarhiz-ium anisopliae Spores Photo by Svetlana Y. Gouli University of Vermont

Figure 17c. Verticillium lecanii Spores Photo by Svetlana Y. Gouli University of Vermont

Figure 18. The density (#/cm) of HWA sistens in field trials in 2003, 4 weeks after treatment with insect-killing fungi at 1 x 10⁸ sp/ml. *Means significantly different from blank control (GLM-ANOVA and Dunnett's). There were no significant differences among pre-treatment counts.

Non-target Effect on HWA Predators

The management of a pest using biologically based strategies often includes more than one component in the management system. If insect-killing fungi are to be deployed in conjunction with predators it is important that these organisms be mutually compatible. During 2001, a petri dish assay system was developed at ERL that provides for both contact and residual exposure of S. tsugae to entomopathogenic fungi. Building on the assay development, laboratory testing for non-target effects against S. tsugae was completed during 2002. Evaluations were done in the forest in 2003 and several fungal isolates were exposed to S. tsugae in sleeve cages on HWA infested hemlock branches similar to those in Figure 11 (page 10). Adult S. tsugae for these trials were provided by the Phillip Alampi Beneficial Insect Lab, New Jersey Department of Agriculture.

Figure 19. Outgrowth of Verticillium lecanii
from an hemlock woolly adelgid cadaver
Photo by Svetlana Y. Gouli, University of Vermont

In the laboratory assay, a concentration of 2 x 108 conidia per milliliter – twice the current field application rate – did not cause any significant decrease in survival rate for S. tsugae for any of the fungi examined, and none of the treated S. tsugae showed signs of fungal outgrowth. However, on day 15

Figure 20. The survival of Sasajiscymnus tsugae in the forests after exposure to insect-killing fungi. *Means significantly different from blank control

Further replicates of the field study are needed, and should include tests against Scymnus spp., Laricobius spp., and other predators being reared for release against HWA.

Acknowledgments

University of Vermont Entomology Research Laboratory (ERL) scientists V. and S. Gouli, M. Brownbridge, W. Reid, and others played an integral role in the development of insect-killing fungi for biological control of HWA.

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