Development of Diagnostic Technology for Monitoring Pyrethroid Resistance Mechanisms in the Tobacco Budworm, Heliothis Virescens (F.).

Abstract

Metabolic resistance was studied in the tobacco budworm, Heliothis virescens (F.) using pyrethroid analogs, insecticide synergists, and an in vitro assay with $\sp{14}$C-cypermethrin. In initial studies, the phenoxybenzyl moiety of conventional pyrethroid, a major site of oxidative metabolism in resistant H. virescens, was replaced with P450 monooxygenase-inhibiting or oxidatively blocked groups. Isomers (1R/1S, cis/trans) of the resulting chrysanthemates were separated and tested as insecticides against pyrethroid-susceptible (LSU) and -resistant H. virescens (Pyr-R). A number of compounds with pentafluorophenyl (PFP), methylenedioxyphenyl (MDP), and propargyloxyphenyl (PP) groups were insecticidal and activity was dependent on both geometric and stereochemical configuration of the acid moiety. Both trans and cis isomers of 1R-fenfluthrin, which contains a PFP group, suppressed resistance to cypermethrin in Pyr-R insects, confirming that oxidative metabolism of the phenoxybenzyl moiety is a major mechanism of resistance in this strain. Of the MDP compounds, 1R, trans and cis isomers were toxic and partially suppressed resistance in Pyr-R larvae. Similarly, both trans and cis isomers of $\alpha S,1R$ PP-containing compounds were insecticidal. In a second approach, studies with pyrethroid synergists showed the importance of enhanced metabolism to pyrethroid-resistance in Pyr-R larvae. Among four P450 monooxygenase inhibitors tested, the propynyl ether, 1, 2, 4-trichloro-3(2-propynyloxy)benzene, was the most effective synergist for cypermethrin toxicity, whereas synergism with piperonyl butoxide (PBO) was moderate. An esterase inhibitor, S,S,S-tributylphosphorotrithioate, also was active as a synergist of cypermethrin toxicity. Non-toxic isomers of pyrethroid analogs were tested as synergists, and only $\alpha R,1R,cis$-MDP- and PP-containing compounds significantly enhanced cypermethrin toxicity. Finally, the enzymes involved in cypermethrin metabolism and resistance in Pyr-R insects were investigated using an in vitro assay. Studies with $\sp{14}$C-cypermethrin revealed that both P450 monooxygenases and esterases were associated with pyrethroid resistance in Pyr-R larvae. HO-cypermethrin, a monooxygenase product, and phenoxybenzoic acid, a product of esterase-mediated hydrolysis, were identified as the two major metabolites. Compared with the LSU strain, field-collected MRS-June and -August H. virescens showed very high levels of the hydrolytic metabolite as well as moderate levels of the oxidative product. Inhibition studies indicated that the monooxygenase inhibitor, PBO, also inhibits esterases.