Assessment and development of bioassay methods for monitoring miticide resistance in spider mites (Tetranychidae)
Abstract
The principal aim of this study was to evaluate bioassay techniques and to determine suitable procedures for detection and monitoring miticide resistance in spider mites with special emphasis on the miticide propargite.
Several bioassay methods were evaluated using propargite (Omite 30% wettable powder (WP)) and fenbutatin oxide (Torque 50% (WP) and 55% suspension concentrate (SC)) with twospotted spider mite (TSM) and European red mite (ERM) to document their utility and reliability or accuracy in lethal concentration (LC) estimates. For each method, two post-treatment exposure periods and mortality criteria were used. Post-treatment exposure period and mortality criteria had a significant influence on the reliability of LC estimates for all tested miticides with all bioassay methods. Twenty four hour (h) post-treatment exposure was found to be the most suitable for the slide dip and Petri dish methods while 48h was the most appropriate for the leaf disc methods. Scoring moribund mites as dead was the most satisfactory criterion for ensuring that bioassays were as simple and reliable as possible. The Petri dish residue- Potter tower method (PDR-PT) estimated the responses of TSM and ERM to propargite with high reliability. The same method did not result in similar reliability with both formulations of fenbutatin oxide. Significant mite run-off occurred with the leaf disc methods, and because of run-off the reliability of these methods cannot be fully known. The slide dip method produced poorly estimated LC values for propargite (WP) and fenbutatin oxide (WP), while the same method produced higher reliability in LC estimates for fenbutatin oxide (SC) than the PDR-PT method. The toxicity of candidate miticides was found to be method- and species-dependent, and LC values between methods within species were significantly different.
Other than the bioassay method, the reliability or precision of LC estimates was found to be influenced by test design (placement of concentrations) and sample size (number of individuals tested for a whole bioassay) employed in a bioassay. Five test designs in combination with five different total sample sizes were investigated in this study using the PDR-PT method with propargite against TSM and ERM. Of the five designs tested, the symmetric-5 design (where concentrations were placed equidistantly on the log scale between the response level of 5-95% mortality) yielded more precise estimates of both the LC₅₀ and LC₉₀ than other designs. With this design, the minimum total sample size requirement was 360, but 480 was found to be more reliable. With a total sample size of ≥600, all designs were equally effective.
The influence of several environmental, operational and biological factors on the toxicity of propargite to TSM and ERM using the PDR-PT method was also investigated. Propargite toxicity was found to be significantly positively correlated to temperature over a range of 10-30° C. Propargite toxicity was also positively correlated to spray volume. When different age groups of adult female TSM were exposed to propargite, toxicity was inversely correlated to adult age. The responses of field collected ERM to propargite varied month by month in an irregular pattern during a growing season (December to April).
The responses of spider mites from one test to another did not vary significantly over time when tests were conducted with a reasonable number of individuals (≥80/concentration) on each occasion. Propargite residues persisted in pre-sprayed dishes for ≥30 days when stored at 4 ± 1°C and remained consistently toxic to mites throughout this period. Propargite did not induce any detectable vapour-activated toxicity in closed Petri dishes. The results of laboratory based Petri dish bioassays were poorly correlated with those obtained under semi-field conditions. Limitations of using laboratory results to determine an effective field application rate of propargite are discussed.
Propargite resistance in numerous TSM and ERM strains was determined by using discriminating concentration (DC) and multiple concentration test techniques. Since the main difficulty with the DC technique was how to determine the appropriate concentration with which to differentiate between resistant and susceptible strains, several ways of selecting a DC were examined using the PDR-PT method. The selected DC (0.105% AI for TSM and 0.045% AI for ERM) identified levels of propargite resistance in different TSM and ERM strains as reliably as did the multiple concentration test. In the case of low resistance, the results of the DC tests were slightly different than those obtained from the multiple concentration tests, but for medium and high levels of resistance both techniques were equally effective. Considering the time involvement and the number of test individuals required for a multiple concentration bioassay, the DC technique would be a cost-effective approach for resistance monitoring. Despite the long history of propargite use in New Zealand, resistance in TSM-and ERM was found to occur sporadically at some locations.
To investigate the dynamics of resistance, a propargite resistant TSM colony was established in the laboratory. Propargite resistance was found to develop fairly rapidly (after three months; one selection in each month) but it did not decline in the absence of selection pressure over six months (approximately 12 generations).
