1. DWFP Toxicology Research to Develop Insecticides for Vector Control Julia W. Pridgeon, PhD Toxicologist/Molecular Biologist 02-14-07 Mosquito and Fly Research Unit Center for Medical, Agricultural, and Veterinary Entomology USDA-ARS, 1600 SW 23rd Drive Gainesville, FL 32608

2. Research Objective To discover and develop effective chemical and molecular insecticides for vector control

3. Military Relevance ♣ In military units, arthropod-borne diseases continue to be serious threats to deployed troops ♣ Chemical pesticides are the most effective way to control disease vectors, however, few products are available for military use ♣ Due to the nature of military operations, pesticides that can quickly and effectively eliminate arthropod vectors are highly desirable

4. Approach for the Development of Effective Chemical Insecticides Registered Pesticides Natural Plant Chemicals Plant Extracts Designer Chemicals USDA-ARS- NPURU USDA-ARS- MFRU NPURU & UFIRAC High-Throughput Screen to find highly toxic chemical (Topical application; Aedes aegypti model) Small Chamber Test and Field Evaluation

5. Topical Application EasyAccurate Small AmountFast

6. Results of Studies on Chemical Insecticides Registered Pesticides Natural Plant Chemicals Plant Extracts 33 Designer Chemicals USDA-ARS- NPURU USDA-ARS- MFRU NPURU & UF IRAC Pesticide LD 50 (ug) Fipronil 1.3  10 -6 Permethrin 1.4  10 -4 Spinosad 2.5  10 -3 Bifenzate 4.25 19 26 AT-05-27-2 (LD 50  0.3ug) 123-1 (LD 50 = 0.4ug) Bacopa monnieri (Water hyssop) Methanol Extract LD 50  125ug 33 LD 50 =0.8ug LD 50 =2.1ug LD 50 =14.72ug

7. Summary on Studies of Chemical Insecticides 1)Screened 19 registered pesticides; several showed high insecticidal activity against Aedes aegypti 2)Screened 26 different natural plant chemicals (NPURU); two had relatively high toxicities

8. 3) Screened 33 different plant extracts (MFRU), some had higher toxicity against mosquitoes than others 4) Screened 33 piperidines with different structures (UF and NPURU), 2-ethyl piperidines showed the highest toxicities against mosquito (in press in Journal of Medical Entomology) Summary on Studies of Chemical Insecticides

9. Future Studies on Chemical Pesticides 1)Evaluate the toxicities of registered pesticides against different species of mosquitoes and other disease vectors 2)Select the best pesticide candidates for further evaluation in small chamber tests and field studies 3)Manipulate the structures of plant chemicals and evaluate their toxicities

10. 4) Studies on toxicity of plant extracts will be continued (additional information by Dr. Quinn) 5) For structure-activity analysis, different classes of chemicals will be evaluated and the best ones chosen for further structure manipulation Future Studies on Chemical Pesticides

11. Approach for the Development of Molecular Pesticides (Aedes aegypti model) Target Essential Pathways Programmed Cell Death, Mitochondria Pathway, etc Molecular Cloning (DNA, RNA, PCR) Vector, Carrier, Formulation (Aedes aegypti model) Bioassays Molecular Pesticide Construction

12. ControlMolecular Pesticide 1 Molecular Pesticide 2Molecular Pesticide 3 Cell Assay Evaluation of Molecular Pesticides

13. Adult Assay Evaluation of Molecular Pesticides ControlMolecular Pesticides

14. 1)Cell and adult assays have demonstrated that critical pathways can be targeted to develop molecular pesticides 2)Using Aedes aegypti as a model organism, molecular pesticides are effective and fast- killing (within 6 hours) Summary Development of Molecular Pesticides

15. 1)Identify new critical pathways to target 2)Produce additional molecular pesticides and evaluate their efficacy 3)Select best molecular pesticides and evaluate production and application methods (formulation, carriers, etc) Future Studies Development of Molecular Pesticides

16. Overall Summary 1)Screened known registered pesticides; several showed high insecticidal activity 2)Screened different natural plant chemicals (NPURU), two had relatively high toxicities against mosquitoes

17. Overall Summary (Cont’d) 3) Screened 33 piperidines (UF & NPURU) with different structures, 2-ethyl piperidine showed the highest toxicity (in press in JME) 4) Cell and adult assays have demonstrated that critical pathways can be targeted to develop molecular pesticides (patent pending) 5) Using Aedes aegypti as a model organism, molecular pesticides are effective and fast- killing (within 6 hours)

18. Acknowledgments Dr. James J. Becnel Dr. Gary G. Clark Dr. Sandra Allan Dr. Dan Kline Dr. Ulrich Bernier Dr. Brian Quinn Dr. Chris Geden Dr. Roberto Pereira Dr. Jerry Hogsette Dr. Ken Linthicum Dr. Liming Zhao Heather Furlong Greg Allen Nathan Newlon Joyce Urban Mosquito and Fly Research Unit, USDA-ARS-CMAVE Dr. Kumudini Meepagala Natural Products Utilization Research Unit, USDA-ARS Dr. Alan Katritzky Department of Chemistry, University of Florida Dr. Daniel Strickman National Program Leader, Program 104, USDA-ARS Dr. Graham White Department of Entomology, University of Florida Deployed War-Fighter Protection Research Program