Characterizing Molecular Mechanisms of Imidacloprid Resistance in Select Populations of Leptinotarsa decemlineata in the Central Sands Region of Wisconsin

Abstract

The Colorado potato beetle, Leptinotarsa decemlineata (Say), is a major agricultural pest in the Central Sands region of Wisconsin. Imidacloprid, a neonicotinoid insecticide, has commonly been used for control of L. decemlineata since its registration in 1995. In the last 10 years, many field populations of L. decemlineata have begun to show increasing imidacloprid resistance. We studied resistance phenotype as a phenomenon that reduces neonicotinoid efficacy and has practical consequences for potato pest management. Although we have not observed complete field failure following the use of these products, multiple studies have demonstrated that the lethal concentration to kill 50% of the test organisms (LC50) in different field populations of L. decemlineata varies greatly which may suggest that resistance of L. decemlineata is heritable and involves genetic changes. An important challenge in understanding resistance is assessing the genetic mechanisms associated with resistance and classifying up-regulated genes that may be involved in combating an insecticide insult. In this study we uncovered trends in imidacloprid phenotypic response that have developed in the region by estimating the LC50 values among different field populations against a range of imidacloprid doses. The LC50 values collected in 2008–2011, and more recently in 2013 and 2014, show that some field locations remain susceptible to imidacloprid, while nearby fields (<100km) have developed high levels of resistance. We also sought to uncover potential mechanisms of resistance at each field location. We compiled a transcriptome for populations, characterized as phenotypically ‘susceptible’ and ‘resistant’, by isolating mRNA from adult beetles and analyzing gene expression level differences. Strong differences were observed in constituently up and down-regulated genes among different field populations. Most significantly, the up-regulation of 3 cytochrome p450s and a glutathione synthetase related protein in multiple resistant populations provide a mechanistic explanation of resistance evolution in L. decemlineata.

Temporal patterns of imidacloprid resistance throughout a growing season in Leptinotarsa decemlineata populations

“…However, there is significant variation in the estimated confidence intervals (CIs), potentially due to the differing sample sizes of adult beetles included in these assays. We also noted that field populations inherently contain a heterogeneous mixture of both resistant and susceptible individuals and variance is frequently high . Despite significant variation in estimated CIs, several time points in the first generation remain statistically more susceptible than their second‐generation counterparts.…”

“…More recently (1995), the registration and introduction of the neonicotinyl insecticide class (IRAC Classification, Group 4A, nicotinic acetylcholine receptor (nAChR) agonists) has resulted in the use of active ingredients, which include, but are not limited to, imidacloprid, thiamethoxam, clothianadin, and dinotefuran . Since the initial introduction of this insecticide class in the mid‐1990s, populations of L. decemlineata have steadily developed resistance, but it remains the principal insecticidal tool used for crop protection in potato …”

“…Recent investigations have examined transcriptomic data to classify possible mechanisms of pesticide resistance in L. decemlineata . These studies examined transcript abundance in relation to insecticide‐resistant populations independent of collection time, with the goal of classifying over‐expressed transcripts and contigs in resistant populations.…”

“…We hypothesize that at time points of increased imidacloprid resistance transcripts that encode for known mechanisms of resistance, such as cytochrome p450s and glutathione S ‐transferases, will be expressed with increased abundance. Furthermore, these peaks in resistance and the associated upregulated transcripts are known to differ between resistant and susceptible populations . Uncovering up‐regulated transcripts provides important new information on how this species combats insecticide inputs and could lead to improvements in our ability to manage resistant populations of this damaging pest.…”

Agricultural fungicides inadvertently influence the fitness of Colorado potato beetles, Leptinotarsa decemlineata, and their susceptibility to insecticides

“…Beetles in this population remain highly susceptible to imidacloprid 8,16 . Healthy adult beetles were hand-collected from the canopy of potato plants, placed in plastic containers and returned to the University of Wisconsin-Madison.…”