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Title: A quantitative in vitro assay for chemical mosquito-deterrent activity without human blood cells

item Klun, Jerome
item Kramer, Matthew
item Zhang, Aijun
item WANG, S.
item DEBBOUN, M.

Submitted to: Journal of the American Mosquito Control Association
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/28/2008
Publication Date: 12/15/2008
Citation: Klun, J.A., Kramer, M.H., Zhang, A., Wang, S., Debboun, M. 2008. A quantitative in vitro assay for chemical mosquito-deterrent activity without human blood cells. Journal of the American Mosquito Control Association. 24(4): 508-512.

Interpretive Summary: Serious diseases are transmitted to humans by blood-feeding mosquitoes. There is a need to discover new effective mosquito bite-deterrent compounds for protection of humans against the bites. Previously, we developed a blood-feeding membrane system, using expired human blood from blood banks, that mosquitoes fed upon as if it were a human being. The blood-feeding system was used to screen for candidate mosquito biting-deterrent chemicals. Using the system, we discovered several new effective candidate human use personal-protection compounds. However, the use of blood was difficult because it was not always available from blood banks, use of it posed a human pathogen health risk and disposal of used blood (a biohazard) was complicated. To avoid these complications, we developed a water solution of salts that could be used instead of blood in the chemical screening process. This makes the process simpler and more efficient for discovery new chemicals to protect humans against pathogens carried by mosquitoes.

Technical Abstract: We report that an aqueous solution containing 10-3 M adenosine triphosphate (ATP) and citrate-phosphate-dextrose-adenine (CPDA-1) can effectively replace transfusable human red blood cells in an in vitro K&D bioassay system for evaluating chemicals for mosquito feeding- deterrent activity, using either Aedes aegypti or Anopheles stephensi. These species fed with similar avidity through collagen membrane covering aqueous 10-3 M ATP plus CPDA-1 or red blood cells in CPDA-1 supplemented with ATP. In a second experiment, we evaluated of the feeding-deterrent activity of Deet and a newly discovered natural product chemical, (-)-isolongifolenone, against these two mosquito species. We found that the feeding-deterrent efficacy of the two chemicals was similar whether the feeding stimulant was red blood cells supplemented with ATP or ATP alone with CPDA-1. Since the use of human red blood cells in bioassays raises important health and logistic issues, aqueous ATP with CPDA-1 is a reasonable alternative to human blood cells for routine in vitro chemical screening.