Behavioral Effects of the Entomopathogenic Fungus,
Entomophthora muscae  on its host Musca domestica :
Postural Control of Dying Hosts and
Gated Pattern of Mortality

S.B. Krasnoff*, D.W. Watson**, D.M. Gibson*, and E.C. Kwan*
*Plant Protection Research Unit
USDA, ARS
Ithaca, NY 14853
**Department of Entomology
Cornell University
Ithaca, NY 14853

Abstract: House flies, Musca domestica  L., infected with the entomopathogenic fungus Entomophthora muscae  typically die in elevated positions, with the proboscis extended and securely fastened to the substrate, the legs spread, the abdomen angled away from the substrate, and the wings raised above the thorax. The behavior of dying diseased flies was recorded on videotape and analyzed. Four events occurring before or shortly after the death were identified, the last locomotory movement (LM), the last extension of the proboscis to the substrate (PD), the start of upward wing movement (WS) and the end of upward wing movement (WU). Sequence progression was unidirectional and highly stereotyped. The median elapsed time between WS and WU was 15 minutes. The median time interval between the LM and WU was 1.25 h. The mortality of diseased flies exhibited a distinct diel periodicity under a variety of natural and artificial LD regimes with most flies dying 0-5 hours before the onset of darkness. The mortality of flies held in the dark from the time of exposure until just before death did no exhibit rhythmicity; however, a circadian periodicity of mortality was observed in a population of infected flies exposed to a 12:12 LD photoperiod for three days following infection and then held in continuous darkness. These findings indicate that mortality in E. muscae -diseased flies is a gated phenomenon governed by a biological clock that, it is argued, is most probably a property of the fungus. J. Insect Physiol.(1995) 41:895-903.

Evaluation of Two Biocontrol Agents
for Prevention of Gray Mold
in Pruning Wounds of Greenhouse Tomatoes

J. S. Lamboy, A. C. Cobb, and H. R. Dillard
Department of Plant Pathology
NYSAES, Cornell University
Geneva, NY 14456

Abstract: One of the most difficult diseases to control on greenhouse tomatoes is gray mold, caused by the fungus Botrytis cinerea.  Traditional methods for control of B. cinerea  on greenhouse tomatoes integrate chemical fungicide sprays and cultural practices. The objective of this project was to determine whether preventive measures with Mycostop, a registered formulation of Streptomyces griseoviridis,  and T22 for the Greenhouse (a wettable formulation of Trichoderma harzianum ) could reduce gray mold infection in pruning wounds of tomato stems. A fractional factorial experiment was designed to evaluate the efficacy of biocontrol agents in preventing infection by B. cinerea.  The two biological control organisms were applied as soil drenches and by spraying on pruning wounds when the axial shoots were removed. The plants were inoculated with a B. cinerea  spore suspension and held in a mist chamber in the greenhouse for two weeks after inoculation. Disease incidence and severity were assessed. This work is part of a larger study in a pesticide-free commercial greenhouse (sponsored by a SARE/ACE grant), in which the results of preventive measures will be monitored until the end of tomato harvest.

Impact of Fungicides and Insecticides on Anagrus epos, parasitoid of Erythroneura leafhoppers

T.E. Martinson and & L. Williams III
Department of Entomology
NYSAES, Cornell University
Geneva, NY 14456

Abstract: Toxicity of fungicides and insecticides used in grape production to Anagrus epos Girault (Hymenoptera: Mymaridae), parasitoid of Erythroneura leafhoppers infesting grapes, was assessed through laboratory and field bioassays. Field-equivalent rates of fungicides were relatively nontoxic to A. epos adults in laboratory bioassays. In bioassays with field-weathered residues, sulfur (9600 ppm) caused elevated mortality of Anagrus adults for 14 to 21 d post treatment. Residues of microencapsulated methyl parathion (600 and 1200 ppm) remained toxic up to 43 d post-treatment. Duration of toxicity of carbaryl to adults was rate dependent, and ranged from >43 d (4800 ppm) to 14 d (1200 ppm), suggesting that the use of reduced rates of carbaryl may reduce disruption of biological control by A. epos in vineyards.

Biological Control of Dicot Weeds in Turf

J.C. Neal
Department of Floriculture and Ornamental Horticulture
Cornell University
Ithaca, NY 14853

Abstract: Herbicides are the predominant weed control method in agricultural production and in the United States account for over 60% of all pesticides applied. In turfgrass areas, herbicide use is estimated to be over 170,000 metric tonnes annually (1991 figures), and may account for up to 90% of the pesticides used on a particular site. Despite the volume of herbicides used, there has been very little research on alternatives to herbicides in turfgrass systems. In collaboration with researchers at Montana State University, University of Minnesota, and several corporations, we are evaluating fungal and bacterial pathogens of dicot weeds commonly infesting turfgrass areas. Two pathogens in particular have shown promise: Sclerotinia sclerotiorum and Pseudomonas syringae pv. tagetis (PST). Both pathogens have relatively broad host ranges on dicot species but are generally not pathogenic to grasses. To restrict the potential for off-target movement, auxotrophic mutants of S. sclerotiorum were evaluated. When applied as mycelial pellets, S. sclerotiorum provided up to 60% control of white clover (Trifolium repens) and thymeleaf speedwell (Veronica serpyllifolia). Dandelion (Taraxacum officinale) was not controlled. PST is a pathogen of many species in the asteraceae. Its efficacy on thistles (Cirsium spp.) and ragweed (Ambrosia artemisiifolia) are well established. Current tests are evaluating PST virulence on numerous aster weeds common to turfgrass areas including dandelion.

Compost-induced Changes in Soil Microbial Communities
Affecting the Suppression of Soilborne Plant Diseases

E. B. Nelson and C.M Craft
Department of Plant Pathology
Cornell University
Ithaca, NY 14853

Abstract: Studies were initiated to establish relationships between the composition and function of microbial communities developing in soils receiving compost amendments and the evolution of disease-suppressive soil properties. Microbial biomass, activity, and functional diversity were detemined over the course of a growing season in turfgrass soils receiving disease- suppressive and non-suppressive compost amendments. Microbial activity (as measured by the hydrolysis of fluorescein diacetate) steadily increased from levels on 30 May to those on 7 August and then leveled off or declined thereafter. The levels of microbial activity in soil amended with some composts were considerably greater during July and August than those in soils treated with other amendments. These general increases in microbial activity were not reflected in plate count populations of bacteria, fungi, and actinomycetes. Functional diversity of microbial communities differed with the type of compost amendment. Metabolic profiles of microbial communities, based on the utilization of a suite of 95 different carbon sources, revealed qualitative differences among microbial communities in soils receiving different compost treatments. These profiles not only differed according to the compost amendment, but they differed temporally. Certain discriminating carbon sources could be identified that were unique to each microbial community and that could be used as signatures for each community. These results reinforce the notion that quantitative measures alone are not adequate to characterize microbial communities in compost-amended soils and that qualitative differences are likely to be key in identifying disease-suppressive properties of certain compost-amended soils.

Are There Characteristics of Natural Enemies Used
in Successful Conservation Biological Control
in Perennial Cropping Systems?

J.P. Nyrop
Department of Entomology
NYSAES, Cornell University
Geneva, NY 14456

Abstract: Conservation biological control entails the manipulation of natural enemies and/or their habitat to enhance their performance as biological control agents. When there are several candidate natural enemies, it may be necessary to ask what species should conservation efforts be directed at. Are there attributes of natural enemies that can be used to aid in this selection process? I compared the dynamics of two phytoseiid mite predators and their prey, the phytophagous European red mite (Panonychus ulmi) that feeds on apple leaves. These two phytoseiid species have very different biologies: One predator, Amblyseius fallacis, is more voracious, is oligophagous and feeds primarily on spider mites, has a faster growth rate, and is more likely to disperse. The other species, Typhlodromus pyri, is polyphagous and feeds on spider mites as well as pollen, is not very voracious, and has a rather weak numerical response. Of the two species, T. pyri is the better biological control agent because it provided consistent (over many years) regulation of prey. Attributes of T. pyri that make it a better biological control agent are higher survival during winter, less propensity to disperse, and the ability to sustain themselves in the absence of spider mites. I suggest that in perennial systems a key process in conservation biological control is maintaining natural enemies in the target habitat. Biological attributes such as being a generalist feeder, having a low propensity to disperse, and being able to survive inhospitable circumstances allow for this process to be realized.

New Associations in Biological Control:
Potential Risks

D. Pimentel
Department of Entomology
Cornell University
Ithaca, NY 14853

Abstract: Nearly 40% of all the cases of successful biological control have been based on new associations between the natural enemy and the pest host. This confirms the effectiveness of introducing a biological control agent from a totally different host and region. The successful introduction of a wasp parasite from a moth host in Virginia to a new and native moth-pest of trees in Columbia, South America illustrates this relationship. The specificity of biocontrol agents that are newly associated with the target pests is similar to biocontrol agents used in classical biocontrol. In addition to the higher rate of successes with new associations compared with classical biological control, the new association approach has the capacity to control native pests which make up 60% t0 80% of all pests. The classical approach does not have this advantage. Environmental problems have not been observed with the introduction of a newly associated biocontrol agent. However, related to classical biological control and old associations, one clear case of an environmental problem occurred when the lace bug was introduced from Mexico into east Africa to control the lantana weed. Ultimately, the bug moved from lantana to attack the sesame crop growing in east Africa. Thus, new associations offer many opportunities for successful biological control with no or minimal potential risks.

Performance of a Leaf-beetle on Different
North American Populations of Purple Loosestrife

M. Schat and B. Blossey
Department of Natural Resources
Cornell University
Ithaca NY, 14850

Abstract: The success of a biological weed control program depends on the ability of the control agent to develop on different host genotypes. We studied the peformance of Galerucella calmariensis, a chrysomelid beetle, on three different North American populations of Lythrum salicaria. Experimental plants were obtained from New York, Minnesota, and Oregon and were grown under identical conditions in Ithaca, New York. We studied the impact and survival of G. calmariensis at three herbivore densities. Plant growth and biomass allocation patterns were significantly different between the tested populations. Survival rates of the leaf feeder (egg to adult) were not significantly different between plant origin or herbivore densities. With increasing densities, herbivory caused significant reductions in plant height, leaf, shoot, root, and total plant biomass. There was no significant interaction between herbivory and plant origin. Plants compensated for the loss of photosynthetic tissue by replacing foliage at the expense of root storage, thus herbivory altered the biomass allocation pattern of L. salicaria.

Biological Control of Pieris rapae 
by Arthropod Predators in Cabbage

M.A. Schmaedick¹, A.M. Shelton¹, and M.P. Hoffmann²
¹Department of Entomology
NYSAES, Cornell University
Geneva, NY 14456
²Department of Entomology
Cornell University
Ithaca, NY14850

Abstract: The role of naturally occurring arthropod predators in reducing populations of imported cabbageworm, Pieris rapae,  in cabbage fields was investigated in four 0.2 ha plots at Geneva, New York. Predator exclusion experiments, sticky traps, and laboratory predation assays provided estimates of overall impact of arthropod predators and information on which species may be important as predators of P. rapae . Survivorship from egg to fifth instar on plants from which predators were excluded by screen cages was compared to survivorship on plants in sham cages that allowed arthropod entry but excluded birds and controlled for cage effect. The experiment was run twice in each of two plots. Although mortality from other causes was responsible for most deaths, comparison of the two treatments in each of the four experiments revealed that exposure to arthropod predators reduced survivorship by 51% to 79%. Survivorship curves indicated that most predation occurred before the second instar. The most abundant predatory arthropods caught in sticky traps placed on cabbage plants in adjacent plots were Lygus  sp., Stenolophus comma, Coleomegilla maculata, Hippodamia variegata, Phalangium opilio,  and Anthicus cervinus.  Laboratory predation assays in small arenas showed that S. comma, C. maculata , and P. opilio  fed readily on P. rapae   eggs and first instars, while L. lineolaris  was much less apt to feed and A. cervinus  fed only rarely on these stages. Ongoing work will employ other methods, including immunological assays and direct observations, to determine which species are most effective as predators of P. rapae  in cabbage.

Fabrication of Artificial Eggs for
Biological Control Programs

A. M. Shelton*, H. G. Craighead**, M. P. Hoffmann&+, and S. W. Turner**
*Department of Entomology
NYSAES, Cornell University
Geneva, NY 14456
**Department of Applied Physics
Cornell University
Ithaca, NY 14853
+Department of Entomology
Cornell University
Ithaca, NY 14853

Abstract: The goal of this project is to create prototype 'artificial eggs' which will serve as rearing units for desirable parasitic wasps, such as Trichogramma spp., which can be released in agricultural settings to control pestiferous insects. This technology will ensure an efficient, cost effective way to produce an abundance of high quality parasites which can be used as biological control agents against damaging insect pests on a number of different crops.

We have tested various shapes and materials for artificial eggs and have shown that the artificial eggs are attractive to the wasps. We have also demonstrated that only a hemispherical egg shape is needed, which will make it easier to make a mold to which diet can be added through the opposite half. We have also shown that the eggs become more attractive to the wasps if they are treated with moth scales or a hexane extract from the scales. Our future efforts will foucs on refining the materials needed for the egg shape and structure and putting the components (egg structure, artificial media, chemical stimulants) together into a prototype which will lend itself to mass production.

Field Evaluation of
Heterorhabditis bacteriophora  'Oswego"
for the Control of Alfalfa Snout Beetle

E. J. Shields, K. L. Flanders and J. M. Miller
Department of Entomology
Cornell University
Ithaca, NY 14853

Abstract: The alfalfa snout beetle, Otiorhynchus ligustici  presently infests ca. 200,000 hectares in eight counties of New York state and southeast Ontario, Canada. Otherwise, there are no other known infestations of this European insect in the Western Hemisphere. The presence of large populations (often several million beetles per hectare) and the cryptic behavior of alfalfa snout beetle adults, insecticide applications fail to adequately control invading beetle populations. Infestations of alfalfa snout beetle frequently destroy an entire alfalfa or clover field in a single growing season, thus limiting the effectiveness and the economy of crop rotation. Because alfalfa snout beetle inhabits the soil as larvae, we have investigated the use of entomopathogenic nematodes for the biological control of alfalfa snout beetle infestations. Weevils have shown a consistent and high susceptibility to entomopathogenic nematodes. Surveys of northern NY in 1990 detected a new strain of the entomopathogenic nematode Heterorhabditis bacteriophora  which was subsequently named 'Oswego'. To date, this nematode has persisted for 5 years from a single introduction in research plots established in 1991 in northern NY. Ongoing research has demonstrated that either the persistent strain of H. bacteriophora  (Oswego) or the nonpersistent strain (NC) provide excellent control of alfalfa snout beetle in the field. Only 'Oswego' survives the winter and is present in high numbers the following year.

IR-4 Biopesticide Grants Program

T. D. Spittler* and C. L. Hartman**
*Cornell Analytical Laboratories
NYSAES, Cornell University
Geneva, NY 14456-0462
**IR-4 Headquarters
PO Box 231
NJAES, Rutgers University
New Brunswick, NJ 08903-0231

Abstract: Interregional Research Project No. 4 helps producers obtain registered pesticides for minor crops and limited uses on major crops, including ornamentals. IR-4Ős primary focus is on the collection of field performance data and the analysis of field samples for chemical residues. The majority of its funding is from USDA-CSREES, but it is also supported by USDA-ARS, commodity organizations, and industry. Cooperators on the project include extension personnel, ARS employees, private contractors, and IR-4 funded university faculty. The IR-4 project is coordinated from the Headquarters Office located at Rutgers, with Regional Laboratories at the University of California-Davis, Michigan State University, University of Florida, Cornell University-Geneva and USDA-ARS, Beltsville handling the field trials and chemical analyses. The program is administered by the IR-4 Technical Committee consisting of Headquarters, CSREES, ARS and Administrative Advisors representatives, plus the four Regional Laboratory Directors.

IR-4 also has an active two-part biopesticide program. The first part is the IR-4 Biopesticide Grants Program which funded nine biopesticide research projects in 1995 and ten in 1996. The second part covers biopesticide petition preparation and submission to EPA. Last year, based on IR-4 petitions, EPA granted tolerance exemptions for methyl anthranilate on blueberries, cherries and grapes; for codling moth granulosis virus on apples, pears, walnuts and plums; and for cinnamaldehyde for mushrooms. In addition, an experimental use permit was granted for two microbials used in potting mix and one is pending for use of a non-aflatoxin producing isolate of Aspergillus flavus as a niche competitor in Arizona cotton. Proposals may be submitted to Christina Hartman at any time: review and allocation is at the Spring Technical Committee Meeting.

Management of Plutella xylostella
on Bt-transgenic Plants:
A Model System

J.D. Tang*, A.M. Shelton*, R.T. Roush**,+,
E.D. Earle++, T.D. Metz++,+++,
B.M. Mitchell*, and H.Lam-Collins*
*Department of Entomology
NYSAES, Cornell University
Geneva, NY 14456
**Department of Entomology
Cornell University
Ithaca, NY 14853
+Current address:
Crop Protection, Waite Institute
University of Adelaide
Glen Osmond, South Australia 5064
++Plant Breeding
Cornell University
Ithaca, NY 14853
+++Current address:
Biological Sciences
Campbell University
Buies Creek, NC 27506

Abstract: In greenhouse studies with CryIA(c)-expressing broccoli, we found that refuge size and spatial distribution had pronounced effects on the evolution of resistance in larvae of the diamondback moth. Refuges that were larger, e.g. 20% refuge, and separate, as opposed to a mixed refuge, appreciably slowed the development of resistance. Implications of these results for other crops are discussed.

Entomopathogenic Fungi as
Biocontrol Agents of Pest Insects
of Agricultural Crops

J. D. Vandenberg, F. A. Cantone, M. Ramos,
M. H. Griggs, J A. Altre, J. L. McManus, and J. E. Williams
USDA/ARS
U.S. Plant, Soil & Nutrition Laboratory
Ithaca, NY 14853

Abstract: Our research on fungal biocontrol of insect pests contributes to broad USDA Agricultural Research Service priorities that emphasize environmentally safe pest control methods with minimal health risks. Target insects for this research now include the Russian wheat aphid (Diuraphis noxia), the diamondback moth (Plutella xylostella), and the potato leafhopper (Empoasca fabae). Our choice of these insects was based on local or regional pest impact, broad mandates for control, or likelihood of successful control using fungi. Candidate fungi being actively studied include Beauveria bassiana, Paecilomyces fumosoroseus and Zoophthora radicans. These have been chosen based on their appropriate host ranges, efficacy for the target pests, and potential for large scale deployment in the field. For applied research, we employ more than merely a "spray and count" approach to ensure successful deployment of fungi as viable agents in pest control systems. Our research includes aspects of production, formulation and delivery as well as monitoring of survival, persistence, and establishment of fungi in agricultural environments. In our basic research program we strive to improve the limited fundamental knowledge of infection processes and mechanisms, virulence factors, and host defenses. Through collaboration with appropriate scientists and agencies, we aim to document these features, demonstrate the compatibility of fungi with other pest management practices, and to transfer technology to action agencies, industry and growers for their use.

Isolation of a Fatty Acid Degradation Mutant of
Enterobacter cloacae  and its Role in
Biological Control of Pythium ultimum

K.V. van Dijk and E.B. Nelson
Department of Plant Pathology
Cornell University
Ithaca, NY 14853

Abstract: Sporangia of Pythium ultimum must germinate in order to incite seed and seedling rots. This germination occurs within 0.5-1 hour after exposure to exudates of germinating seeds. In cotton, and possibly other plant species, fatty acids are important germination stimulation molecules, with linoleic acid being the most stimulatory. Enterobacter cloacae strain EcCT-501, an effective biological seed protectant, is capable of inactivating the stimulatory activity of seed exudates and linoleic acid. Using TnphoA mutagenesis, several mutants have been obtained that are incapable of growing on linoleic acid as a sole carbon source. One of these mutants, Ec31, has an insertion in fadB, encoding part of a multi-enzyme complex involved in the b-oxidation of fatty acids. This mutant fails to inactivate the germination stimulatory activity of cotton seed exudate and linoleic acid, and has reduced abilities to protect cotton from seed and seedling rots, and to rapidly. In addition, Ec31 removes linoleic acid from solution at a much slower rate than wildtype E. cloacae. Our data suggest that metabolism of exudate fatty acids is an important mechanism used by E. cloacae to protect seeds and seedlings from Pythium damping off.

A Novel Insect Protein and its Interaction
With a Baculovirus Enhancing Protein

P. Wang*,** and Robert R. Granados**
*Department of Entomology
Cornell University
Ithaca, NY 14850
**Boyce Thompson Institute
Cornell University
Ithaca, NY 14853

Abstract: Insect midgut peritrophic membranes are a significant biological barrier to microbial pathogens. We report the identification and characterization of a novel insect protein from Trichoplusia ni  peritrophic membranes. Biochemical characteristics of this novel protein indicate that the protein plays protective roles in the insect midgut. Interaction of a baculovirus enhancing protein with this novel protein was also investigated. Biochemical studies and both in vitro and in vivo peritrophic membrane assays suggest the the interaction of the enhancing protein with the novel protein leads to increased viral infection of the insect host.

Resistance to Plant Viruses Induced by Incorporation
of a Viral Replicase Gene into the Host Genome

W.M. Wintermantel, N. Banerjee, and M. Zaitlin
Department of Plant Pathology
Cornell University
Ithaca, NY 14853

Abstract: Plant viruses contain genes which encode proteins allowing them to infect and replicate in their host plants. One or more of their genes encode proteins known as replicases, which are associated with virus reproduction. When a replicase gene is cloned from the virus and inserted into the genome of a host plant, the plant often becomes resistant to infection by the virus from which the gene was derived. This technique, known as replicase-mediated resistance, has been utilized to create resistance to a number of plant viruses in many different host plants. Resistance is specific only to the the virus from which the transgene is derived and to closely related viruses, but plants are resistant to inoculation with high levels of either intact virus particles or viral nucleic acid. Resistance is characterized by decreased virus replication, and in some cases by a reduced ability to be transported from cell-to-cell or through the vascular system of the host.