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Horizontal transmission of fipronil in the eastern subterranean termite from Indiana

Dissertation
Author: Jody Michelle Green
Abstract:
Fipronil was tested against the eastern subterranean termite, Reticulitermes flavipes (Kollar) to determine toxicological effects of various exposure methods and possible mechanisms of horizontal transmission between topically-treated donors and untreated recipient nestmates. In laboratory assays, individual termite workers were treated topically with acute doses of fipronil which resulted in a 24 and 48 h LD50 of 1.2 and 0.76 ng per insect, respectively. Topical concentrations of fipronil at doses 2.5 and 5.0 ng per insect resulted in LT50 s of 10.3 and 3 h, respectively. Symptomatic termites exhibited uncoordinated movement, tightly curled tarsi, and paralysis, which always lead to death. Reaction to fipronil-treated paper resulted in similar, yet delayed responses for termites allowed to walk on 12 concentrations. After 24 h exposure, LC 50 and LC90 were 3.5 and 7 ppm (wt AI: wt filter paper), respectively. After 48 h of continuous exposure to treated filter paper, any concentration greater than 2 ppm resulted in 100% mortality. Termites suffered quick and complete mortality when exposed to a 1:1 mixture of fipronil-treated sand: vermiculite substrate. No repellency was observed as termites readily tunneled into 4 cm of each concentration, which ranged from 0.5 - 75 ppm (wt AI: wt substrate). Substrate concentrations greater than 6 ppm achieved 100 % mortality within 24 h. As expected, there was a dose-response effect, where higher concentrations of fipronil resulted in faster onset of symptoms and rapid moribundity and mortality. Fipronil and three of its metabolites were tested with topical applications. Fipronil and fipronil-desulfinyl had identical toxicological effects at a dose of 10 ng per insect. Fipronil-sulfide had the greatest mortality of the four compounds tested at 3 and 6 h observation time points. Fipronil-sulfone was the slowest to result in symptoms of poisoning and mortality when compared to the other three compounds. Using GC analysis, fipronil was found to metabolize via biological pathways in termite bodies to fipronil-sulfone. In tests with fipronil and fipronil-sulfone at low doses (1.0 - 5.0 ng per insect), LD 50 s were similar and LD-P graphs had similar slopes. Topical LD50 s were 1.3 and 2.3 ng per termite for fipronil and fipronil-sulfone, respectively. Horizontal transfer studies, using 14 C-radiolabeled fipronil, were performed to monitor mortality of recipients and the distribution of fipronil within a closed system. Blue-stained donors were treated topically with 5 ng per insect and allowed to interact with 15 untreated, unstained recipients. Donor-Recipient ratios were 1:15, 1:5, and 1:3. Within 24 h, donors were able to transfer more than 50% of fipronil to the immediate environment. For all treatments, approximately 10 % fipronil was recovered from the filter paper, 25 -40% remained in the donor, and 50 - 66% was recovered from recipients. Increasing the number of donors treated with 5 ng per termite increased recipient mortality, and increasing the surface are of the Petri-dish arena increased recipient mortality as well. Using GC Analysis fipronil-sulfone was recovered in donors and recipients from transfer studies. Fipronil was transferred from donors to recipients by allogrooming, however, fipronil-sulfone, which has greater water solubility than the parent material, may be transferred among nestmates via trophallaxis.

Graduate School ETD Form 9 (Revised 12/07) PURDUE UNIVERSITY GRADUATE SCHOOL Thesis/Dissertation Acceptance This is to certify that the thesis/dissertation prepared By Entitled For the degree of Is approved by the final examining committee:

Chair

To the best of my knowledge and as understood by the student in the Research Integrity and Copyright Disclaimer (Graduate School Form 20), this thesis/dissertation adheres to the provisions of Purdue University’s “Policy on Integrity in Research” and the use of copyrighted material.

Approved by Major Professor(s): ____________________________________ ____________________________________ Approved by:

Head of the Graduate Program Date Jody Michelle Green Horizontal Transmission Of Fipronil In The Eastern Subterranean Termite FromIndiana Doctor of Philosophy Gary Bennett Alan York Michael Scharf Ronald Turco Gary Bennett Steve Yaninek 05/30/2008

Graduate School Form 20 (Revised 10/07) PURDUE UNIVERSITY GRADUATE SCHOOL Research Integrity and Copyright Disclaimer Title of Thesis/Dissertation: For the degree of ________________________________________________________________ I certify that in the preparation of this thesis, I have observed the provisions of Purdue University Executive Memorandum No. C-22, September 6, 1991, Policy on Integrity in Research.* Further, I certify that this work is free of plagiarism and all materials appearing in this thesis/dissertation have been properly quoted and attributed. I certify that all copyrighted material incorporated into this thesis/dissertation is in compliance with the United States’ copyright law and that I have received written permission from the copyright owners for my use of their work, which is beyond the scope of the law. I agree to indemnify and save harmless Purdue University from any and all claims that may be asserted or that may arise from any copyright violation. ________________________________ Signature of Candidate ________________________________ Date *Located at http://www.purdue.edu/policies/pages/teach_res_outreach/c_22.html Horizontal Transmission Of Fipronil In The Eastern Subterranean Termite FromIndiana Doctor of Philosophy 05/30/2008 Jody Michelle Green

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Full document contains 151 pages
Abstract: Fipronil was tested against the eastern subterranean termite, Reticulitermes flavipes (Kollar) to determine toxicological effects of various exposure methods and possible mechanisms of horizontal transmission between topically-treated donors and untreated recipient nestmates. In laboratory assays, individual termite workers were treated topically with acute doses of fipronil which resulted in a 24 and 48 h LD50 of 1.2 and 0.76 ng per insect, respectively. Topical concentrations of fipronil at doses 2.5 and 5.0 ng per insect resulted in LT50 s of 10.3 and 3 h, respectively. Symptomatic termites exhibited uncoordinated movement, tightly curled tarsi, and paralysis, which always lead to death. Reaction to fipronil-treated paper resulted in similar, yet delayed responses for termites allowed to walk on 12 concentrations. After 24 h exposure, LC 50 and LC90 were 3.5 and 7 ppm (wt AI: wt filter paper), respectively. After 48 h of continuous exposure to treated filter paper, any concentration greater than 2 ppm resulted in 100% mortality. Termites suffered quick and complete mortality when exposed to a 1:1 mixture of fipronil-treated sand: vermiculite substrate. No repellency was observed as termites readily tunneled into 4 cm of each concentration, which ranged from 0.5 - 75 ppm (wt AI: wt substrate). Substrate concentrations greater than 6 ppm achieved 100 % mortality within 24 h. As expected, there was a dose-response effect, where higher concentrations of fipronil resulted in faster onset of symptoms and rapid moribundity and mortality. Fipronil and three of its metabolites were tested with topical applications. Fipronil and fipronil-desulfinyl had identical toxicological effects at a dose of 10 ng per insect. Fipronil-sulfide had the greatest mortality of the four compounds tested at 3 and 6 h observation time points. Fipronil-sulfone was the slowest to result in symptoms of poisoning and mortality when compared to the other three compounds. Using GC analysis, fipronil was found to metabolize via biological pathways in termite bodies to fipronil-sulfone. In tests with fipronil and fipronil-sulfone at low doses (1.0 - 5.0 ng per insect), LD 50 s were similar and LD-P graphs had similar slopes. Topical LD50 s were 1.3 and 2.3 ng per termite for fipronil and fipronil-sulfone, respectively. Horizontal transfer studies, using 14 C-radiolabeled fipronil, were performed to monitor mortality of recipients and the distribution of fipronil within a closed system. Blue-stained donors were treated topically with 5 ng per insect and allowed to interact with 15 untreated, unstained recipients. Donor-Recipient ratios were 1:15, 1:5, and 1:3. Within 24 h, donors were able to transfer more than 50% of fipronil to the immediate environment. For all treatments, approximately 10 % fipronil was recovered from the filter paper, 25 -40% remained in the donor, and 50 - 66% was recovered from recipients. Increasing the number of donors treated with 5 ng per termite increased recipient mortality, and increasing the surface are of the Petri-dish arena increased recipient mortality as well. Using GC Analysis fipronil-sulfone was recovered in donors and recipients from transfer studies. Fipronil was transferred from donors to recipients by allogrooming, however, fipronil-sulfone, which has greater water solubility than the parent material, may be transferred among nestmates via trophallaxis.