Long-term survey and characterization of cyflumetofen resistance in Tetranychus urticae populations from Turkey.

The two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae) is the most economically important mite pest in agricultural areas and chemical acaricides are widely used to control T. urticae populations. Cyflumetofen is a recently introduced acaricide that inhibits the mitochondrial electron transport chain at complex II (succinate dehydrogenase, SDH), which represents the most recently developed mode of action for mite control worldwide. In the present study, started upon the launch of cyflumetofen in Turkey, a five-year survey was performed to monitor cyflumetofen susceptibility in 28 T. urticae populations collected from agricultural fields across the country. The first resistance case that might cause control failure in practical field conditions was uncovered in 2019, three years after the registration of cyflumetofen. In addition, an extremely resistant population (1722-fold resistance) was also detected towards the end of 2019. Cyflumetofen resistance did not decrease in the laboratory after relaxation of selection pressure for over one year in field-collected populations, suggesting the absence of a fitness cost associated with resistance in these populations. Next to phenotypic resistance, metabolic and physiological mechanisms underlying the decreased susceptibility were also investigated. Synergism assays showed the involvement of P450 monooxygenases in cyflumetofen resistance. Downregulation of carboxylesterases as resistance mechanism, is underpinned by the fact that pre-treatment with esterase inhibitor DEF decreased cyflumetofen toxicity in field-collected strains. Furthermore, a novel H258L substitution in the subunit B of complex II was uncovered in a field population. In silico modeling of the new mutation suggested that the mutation might indeed influence toxicity to complex II inhibitors cyenopyrafen and pyflubumide, but most likely not cyflumetofen. However, further studies are needed to uncover the exact role of this mutation in resistance to this new class of complex II inhibitors.

Pirimicarb resistance and associated mechanisms in field-collected and selected populations of Neoseiulus californicus.

The predatory mite Neoseiulus californicus McGregor (Acari: Phytoseiidae) is an important natural enemy of phytophagous mites, and naturally established populations are often found in apple orchards. However, insecticide applications to control pests cause side effects to non-target organisms such as N. californicus. Pirimicarb, a widely used carbamate insecticide in apple orchards, is generally considered a selective aphidicide, however, toxicity to beneficial insects and predatory mites has been reported. Furthermore, the molecular basis for this selectivity, if present in N. californicus, is still largely unknown. In this study, 8 field-collected N. californicus populations were investigated and showed up to 27-fold resistance compared to a susceptible laboratory population. Selection in the laboratory for 5 consecutive generations resulted in a 69-fold pirimicarb resistance. Although there were no significant difference in terms of the acetlycholinesterase (AChE) activity between susceptible and field-collected populations, the selected population exhibited a significantly higher AChE activity. In addition, gene copy number variation of acetylcholinesterase (ace) gene among populations was detected and ranged from 1.6 to 2.1-fold relative to the susceptible population. All field-collected populations, but not the selected population, had a significantly higher ace copy number compared to the susceptible population (t-test, p < 0.05). Molecular analysis of the target-site (AChE) revealed, for the first time, a phenylalanine to tryptophan substition at position 331 in AChE (Torpedo californica numbering), both in field-collected and the selected population, but not in the susceptible population. Last, the selected F5 population consumed significantly more Tetranychusurticae adults than the parental population. Together, the results of this study shed light on the molecular determinants of acaricide selectivity in predatory mites, and will contribute to a better design of an integrated mite management program, including the use of pesticide resistant N. californicus in apple orchards.