Larvae of an invasive scarab increase greenhouse gas emissions from soils and recruit gut mycobiota involved in C and N transformations.

Background: Soil-derived prokaryotic gut communities of the Japanese beetle Popillia japonica Newman (JB) larval gut include heterotrophic, ammonia-oxidizing, and methanogenic microbes potentially capable of promoting greenhouse gas (GHG) emissions. However, no research has directly explored GHG emissions or the eukaryotic microbiota associated with the larval gut of this invasive species. In particular, fungi are frequently associated with the insect gut where they produce digestive enzymes and aid in nutrient acquisition. Using a series of laboratory and field experiments, this study aimed to (1) assess the impact of JB larvae on soil GHG emissions; (2) characterize gut mycobiota associated with these larvae; and (3) examine how soil biological and physicochemical characteristics influence variation in both GHG emissions and the composition of larval gut mycobiota. Methods: Manipulative laboratory experiments consisted of microcosms containing increasing densities of JB larvae alone or in clean (uninfested) soil. Field experiments included 10 locations across Indiana and Wisconsin where gas samples from soils, as well as JB and their associated soil were collected to analyze soil GHG emissions, and mycobiota (ITS survey), respectively. Results: In laboratory trials, emission rates of CO2, CH4, and N2O from infested soil were ≥ 6.3× higher per larva than emissions from JB larvae alone whereas CO2 emission rates from soils previously infested by JB larvae were 1.3× higher than emissions from JB larvae alone. In the field, JB larval density was a significant predictor of CO2 emissions from infested soils, and both CO2 and CH4 emissions were higher in previously infested soils. We found that geographic location had the greatest influence on variation in larval gut mycobiota, although the effects of compartment (i.e., soil, midgut and hindgut) were also significant. There was substantial overlap in the composition and prevalence of the core fungal mycobiota across compartments with prominent fungal taxa being associated with cellulose degradation and prokaryotic methane production/consumption. Soil physicochemical characteristics such as organic matter, cation exchange capacity, sand, and water holding capacity, were also correlated with both soil GHG emission, and fungal a-diversity within the JB larval gut. Conclusions: Results indicate JB larvae promote GHG emissions from the soil directly through metabolic activities, and indirectly by creating soil conditions that favor GHG-associated microbial activity. Fungal communities associated with the JB larval gut are primarily influenced by adaptation to local soils, with many prominent members of that consortium potentially contributing to C and N transformations capable of influencing GHG emissions from infested soil.

Pest population dynamics are related to a continental overwintering gradient.

Overwintering success is an important determinant of arthropod populations that must be considered as climate change continues to influence the spatiotemporal population dynamics of agricultural pests. Using a long-term monitoring database and biologically relevant overwintering zones, we modeled the annual and seasonal population dynamics of a common pest, Helicoverpa zea (Boddie), based on three overwintering suitability zones throughout North America using four decades of soil temperatures: the southern range (able to persist through winter), transitional zone (uncertain overwintering survivorship), and northern limits (unable to survive winter). Our model indicates H. zea population dynamics are hierarchically structured with continental-level effects that are partitioned into three geographic zones. Seasonal populations were initially detected in the southern range, where they experienced multiple large population peaks. All three zones experienced a final peak between late July (southern range) and mid-August to mid-September (transitional zone and northern limits). The southern range expanded by 3% since 1981 and is projected to increase by twofold by 2099 but the areas of other zones are expected to decrease in the future. These changes suggest larger populations may persist at higher latitudes in the future due to reduced low-temperature lethal events during winter. Because H. zea is a highly migratory pest, predicting when populations accumulate in one region can inform synchronous or lagged population development in other regions. We show the value of combining long-term datasets, remotely sensed data, and laboratory findings to inform forecasting of insect pests.

Salmonella enterica changes Macrosteles quadrilineatus feeding behaviors resulting in altered S. enterica distribution on leaves and increased populations.

Hemipteran insects are ubiquitous inhabitants of the phyllosphere. Changes in microbial phyllosphere communities have recently been demonstrated following infestation by Macrosteles quadrilineatus (Aster Leafhopper). Although epiphytic Salmonella enterica populations naturally decline in the phyllosphere of plants, M. quadrilineatus infestation facilitated the growth of the bacterial pathogen populations. Here, we demonstrate that cellular damage by insect stylet penetration results in a localized beneficial niche on the leaf surface, leading to enhanced S. enterica populations. We measured S. enterica populations and colonization patterns on plants infested with Hemipterans with distinct feeding behaviors. M. quadrilineatus infestation resulted in higher solute leakage and significantly greater bacterial populations than plants absent of insects. Following immigration via contaminated irrigation water, the highest populations of S. enterica are naturally found on the tips of tomato leaflets. We discovered M. quadrilineatus feeding preference altered the natural distribution of S. enterica populations, and that the presence of S. enterica altered the distribution of probing attempts. These findings elucidate how cellular damage resulting from insect feeding drives changes in bacterial colonization of the phyllosphere.

Targeted RNA sequencing reveals differential patterns of transcript expression in geographically discrete, insecticide resistant populations of Leptinotarsa decemlineata.

BACKGROUND: The Colorado potato beetle (Leptinotarsa decemlineata Say) is a major agricultural pest of commercial potatoes, partially due to its ability to rapidly develop resistance to multiple insecticide modes of action. Patterns of L. decemlineata insecticide resistance in the contiguous United States have been linked to geographic location and regional management practices. Several previous studies have classified enzymes that are overexpressed following L. decemlineata exposure to commercial pesticides, many of which have been linked to xenobiotic metabolism. Studies have further associated geographic disparities in resistance patterns to cross-resistance driven by fungicide exposure in the East Coast and Midwest. RESULTS: In this study, our objective was to investigate transcript expression of 38 previously classified detoxification enzymes induced by imidacloprid (an insecticide) and chlorothalonil (a fungicide) within five discrete populations of L. decemlineata obtained from areas in the USA representing eastern, midwestern and western production regions. We found unique patterns of transcript expression in different geographic locations, including overexpression of transcripts related to insecticide metabolism within insecticide-resistant populations. CONCLUSION: The results suggest the genetic response of these populations may be partially linked to geographic location and corresponding management practices. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

'Candidatus Phytoplasma asteris' subgroups display distinct disease progression dynamics during the carrot growing season.

Aster Yellows phytoplasma (AYp; 'Candidatus Phytoplasma asteris') is an obligate bacterial pathogen that is the causative agent of multiple diseases in herbaceous plants. While this phytoplasma has been examined in depth for its disease characteristics, knowledge about the spatial and temporal dynamics of pathogen spread is lacking. The phytoplasma is found in plant's phloem and is vectored by leafhoppers (Cicadellidae: Hemiptera), including the aster leafhopper, Macrosteles quadrilineatus Forbes. The aster leafhopper is a migratory insect pest that overwinters in the southern United States, and historical data suggest these insects migrate from southern overwintering locations to northern latitudes annually, transmitting and driving phytoplasma infection rates as they migrate. A more in-depth understanding of the spatial, temporal and genetic determinants of Aster Yellows disease progress will lead to better integrated pest management strategies for Aster Yellows disease control. Carrot, Daucus carota L., plots were established at two planting densities in central Wisconsin and monitored during the 2018 growing season for Aster Yellows disease progression. Symptomatic carrots were sampled and assayed for the presence of the Aster Yellows phytoplasma. Aster Yellows disease progression was determined to be significantly associated with calendar date, crop density, location within the field, and phytoplasma subgroup.

Frankliniella occidentalis facilitate Salmonella enterica survival in the phyllosphere.

The human enteric bacterial pathogen Salmonella enterica causes approximately 1.35 million cases of food borne illnesses annually in the United States. Of these salmonellosis cases, almost half are derived from the consumption of fresh, raw produce. Although epiphytic S. enterica populations naturally decline in the phyllosphere, a subset of phytophagous insects have recently been identified as biological multipliers, consequently facilitating the growth of bacterial populations. We investigated whether tomato leaves with macroscopic feeding damage, caused by infestation of adult Western flower thrips (Frankliniella occidentalis), support higher S. enterica populations. To explore this hypothesis, we assessed S. enterica populations in response to thrips feeding by varying insect density, plant age, and the gender of the insect. As a reference control, direct leaf damage analogous to thrips feeding was also evaluated using directed, hydraulic pressure. In a supplementary set series of experiments, groups of F. occidentalis infested tomato plants were later inoculated with S. enterica to determine how prior insect infestation might influence bacterial survival and persistence. Following an infestation period, leaves visibly damaged by adult F. occidentalis supported significantly higher S. enterica populations and resulted in greater amounts of electrolyte leakage (measured as electrical conductivity) than leaves lacking visible feeding damage. Plant age did not significantly influence S. enterica populations or estimates of electrolyte leakage, independent of initial infestation. Additionally, the gender of the insect did not uniquely influence S. enterica population dynamics. Finally, applications of aggressive water bombardment resulted in more electrolyte leakage than leaves damaged by F. occidentalis, yet supported comparable S. enterica populations. Together, this study indicates that F. occidentalis feeding is one of the many potential biological mechanisms creating a more habitable environment for S. enterica.

Characterization of calcium signaling proteins from the fat body of the Colorado Potato Beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae): Implications for diapause and lipid metabolism.

Calcium (Ca2+) regulates many cellular and physiological processes from development to reproduction. Ca2+ is also an important factor in the metabolism of lipids, the primary energy source used during insect starvation and diapause. Ca2+ signaling proteins bind to Ca2+ and maintain intracellular Ca2+ levels. However, knowledge about Ca2+ signaling proteins is mostly restricted to the model Drosophila melanogaster and the response of Ca2+ signaling genes to starvation or diapause is not known. In this study, we identified three Ca2+ signaling proteins; the primary Ca2+ binding protein Calmodulin (LdCaM), phosphatase Calcineurin B (LdCaNB), and the senescence marker protein Regucalcin (LdRgN), from the fat body of the Colorado Potato Beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae). This insect is a major pest of potato worldwide and overwinters under hibernation diapause as adults while utilizing lipids as the primary energy source. Putative EF-hand domains involved in Ca2+ binding were present in LdCaM, LdCaNB, but absent in LdRgN. LdCaM and LdCaNB were expressed in multiple tissues, while LdRgN was primarily expressed in the fat body. LdCaM was constitutively-expressed throughout larval development and at the adult stage. LdCaNB was primarily expressed in feeding larvae, and LdRgN in both feeding larvae and adults at comparable levels; however, both genes were down-regulated by molting. A response to starvation was observed only for LdRgN. Transcript abundance analysis in the entire body in relation to diapause revealed differential regulation with a general suppression during diapause, and higher mRNA levels in favor of females at post-diapause for LdCaM, and in favor of males at non-diapause for LdCaNB. Fat body-specific transcript abundance was not different between non-diapause and post-diapause for LdCaNB, but both LdCaM and LdRgN were down-regulated in males and both sexes, respectively by post-diapause. Silencing LdCaNB or LdRgN in larvae led to decreased fat content, indicating their involvement in lipid accumulation, while RNAi of LdCaM led to lethality.

Two calcium-binding chaperones from the fat body of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) involved in diapause.

Molecular chaperones are crucial for the correct folding of newly synthesized polypeptides, in particular, under stress conditions. Various studies have revealed the involvement of molecular chaperones, such as heat shock proteins, in diapause maintenance and starvation; however, the role of other chaperones in diapause and starvation relatively is unknown. In the current study, we identified two lectin-type chaperones with calcium affinity, a calreticulin (LdCrT) and a calnexin (LdCnX), that were present in the fat body of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) during diapause. Both proteins possessed an N-globular domain, a P-arm domain, and a highly charged C-terminal domain, while an additional transmembrane domain was present in LdCnX. Phylogenetic analysis revealed distinction at the order level. Both genes were expressed in multiple tissues in larval and adult stages, and constitutively throughout development, though a starvation response was detected only for LdCrT. In females, diapause-related expression analysis in the whole body revealed an upregulation of both genes by post-diapause, but a downregulation by diapause only for LdCrT. By contrast, males revealed no alteration in their diapause-related expression pattern in the entire body for both genes. Fat body-specific expression analysis of both genes in relation to diapause revealed the same expression pattern with no alteration in females and downregulation in males by post-diapause. This study suggests that calcium-binding chaperones play similar and possibly gender-specific roles during diapause.

Degree day models to forecast the seasonal phenology of Drosophila suzukii in tart cherry orchards in the Midwest U.S.

Spotted-wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), is an invasive economic pest of soft-skinned and stone fruit across the globe. Our study establishes both a predictive generalized linear mixed model (GLMM), and a generalized additive mixed model (GAMM) of the dynamic seasonal phenology of D. suzukii based on four years of adult monitoring trap data in Wisconsin tart cherry orchards collected throughout the growing season. The models incorporate year, field site, relative humidity, and degree days (DD); and relate these factors to trap catch. The GLMM estimated a coefficient of 2.21 for DD/1000, meaning for every increment of 1000 DD, trap catch increases by roughly 9 flies. The GAMM generated a curve based on a cubic regression smoothing function of DD which approximates critical DD points of first adult D. suzukii detection at 1276 DD, above average field populations beginning at 2019 DD, and peak activity at 3180 DD. By incorporating four years of comprehensive seasonal phenology data from the same locations, we introduce robust models capable of using DD to forecast changing adult D. suzukii populations in the field leading to the application of more timely and effective management strategies.

Genetic Variation Among Geographically Disparate Isolates of Aster Yellows Phytoplasma in the Contiguous United States.

Aster Yellows phytoplasma (AYp; Candidatus Phytoplasma asteris) is associated with diseases of herbaceous plants, including ornamentals and important commercial vegetable and grain crops. The aster leafhopper (ALH; Macrosteles quadrilineatus Forbes) is the predominant vector of these bacteria, though other leafhopper species can acquire and transmit AYp. Potentially inoculative leafhoppers are reported to overwinter in the southern United States and migrate to northern latitudes in the spring. Examining the genetic similarities and differences in AYp associated with southern and northern populations of ALH may provide insight into the role that migrating ALH play in AYp disease development. To investigate similarities among geographically distinct populations of ALH and characterize the variation in AYp associated within these populations, we identified genetic variations in subgroup designation and the relative proportions of secreted AY-WB proteins from field-collected populations of AYp isolated from ALH from select locations in the southern (Arkansas, Kansas, Oklahoma, and Texas) and the northern United States (Wisconsin) in 2016, 2017, and 2018. Isolated phytoplasma were tested for variation of AYp genotypes, numbers of potentially inoculative (AYp-positive) ALH, and presence of specific AYp virulence (effector) genes. Geographically distinct populations of ALH collected in northern and southern regions were similar in CO1 genotype but carried different proportions of AYp genotypes. While similar AYp strains were detected in geographically distinct locations, the proportion of each genotype varied over time.

Changes in emergence phenology, fatty acid composition, and xenobiotic-metabolizing enzyme expression is associated with increased insecticide resistance in the Colorado potato beetle.

The Colorado potato beetle (Leptinotarsa decemlineata) is a major agricultural pest of solanaceous crops. An effective management strategy employed by agricultural producers to control this pest species is the use of systemic insecticides. Recent emphasis has been placed on the use of neonicotinoid insecticides. Despite efforts to curb resistance development through integrated pest management approaches, resistance to neonicotinoids in L. decemlineata populations continues to increase. One contributing factor may be alterations in insect fatty acids, which have multiple metabolic functions and are associated with the synthesis of xenobiotic-metabolizing enzymes to mitigate the effects of insecticide exposure. In this study, we analyzed the fatty acid composition of L. decemlineata populations collected from an organic production field and from a commercially managed field to determine if fatty acid composition varied between the two populations. We demonstrate that a population of L. decemlineata that has a history of systemic neonicotinoid exposure (commercially managed) has a different lipid composition and differential expression of known metabolic detoxification mechanisms relative to a population that has not been exposed to neonicotinoids (organically managed). The fatty acid data indicated an upregulation of Δ6 desaturase in the commercially managed L. decemlineata population and suggest a role for eicosanoids and associated metabolic enzymes as potential modulators of insecticide resistance. We further observed a pattern of delayed emergence within the commercially managed population compared with the organically managed population. Variations in emergence timing together with specific fatty acid regulation may significantly influence the capacity of L. decemlineata to develop insecticide resistance.

Conjugated linoleic acid as a novel insecticide targeting the agricultural pest Leptinotarsa decemlineata.

The Colorado Potato Beetle, Leptinotarsa decemlineata, is a major agricultural pest of solanaceous crops in the United States. Historically, a multitude of insecticides have been used to control problematic populations. Due to increasing resistance to insecticides, novel compounds and methodologies are warranted for the control of beetle populations. Mixed-isomer conjugated linoleic acid has been studied in-depth for its beneficial properties to mammalian systems. At the same time, studies have demonstrated that conjugated linoleic acid can manipulate fatty acid composition in non-mammalian systems, resulting in embryo mortality. Consequently, experiments were conducted to assess the effects of foliar-applied conjugated linoleic acid on larval growth, embryogenesis, and feeding preference in Colorado potato beetle. Both maternal and deterrent effects of dietary conjugated linoleic acid were assessed. Conjugated linoleic acid demonstrated desirable insecticidal properties, including increased larval mortality, slowed larval development, antifeedant effects, and decreased egg viability after maternal ingestion.

Widespread detections of neonicotinoid contaminants in central Wisconsin groundwater.

Neonicotinoids are a popular and widely-used class of insecticides whose heavy usage rates and purported negative impacts on bees and other beneficial insects has led to questions about their mobility and accumulation in the environment. Neonicotinoid compounds are currently registered for over 140 different crop uses in the United States, with commercial growers continuing to rely heavily on neonicotinoid insecticides for the control of key insect pests through a combination of in-ground and foliar applications. In 2008, the Wisconsin Department of Agriculture, Trade and Consumer Protection (DATCP) began testing for neonicotinoids in groundwater test wells in the state, reporting detections of one or more neonicotinoids in dozens of shallow groundwater test wells. In 2011, similar detection levels were confirmed in several high-capacity overhead center-pivot irrigation systems in central Wisconsin. The current study was initiated to investigate the spatial extent and magnitude of neonicotinoid contamination in groundwater in and around areas of irrigated commercial agriculture in central Wisconsin. From 2013-2015 a total of 317 samples were collected from 91 unique high-capacity irrigation wells and tested for the presence of thiamethoxam (TMX), a neonicotinoid, using enzyme-linked immunosorbent assays. 67% of all samples were positive for TMX at a concentration above the analytical limit of quantification (0.05 µg/L) and 78% of all wells tested positive at least once. Mean detection was 0.28 µg/L, with a maximum detection of 1.67 µg/L. Five wells had at least one detection exceeding 1.00 µg/L. Furthermore, an analysis of the spatial structure of these well detects suggests that contamination profiles vary across the landscape, with differences in mean detection levels observed from landscape (25 km), to farm (5 km), to individual well (500 m) scales. We also provide an update of DATCP's neonicotinoid monitoring in Wisconsin's shallow groundwater test wells and private potable wells for the years 2011-2017.

Transcriptomic analysis reveals similarities in genetic activation of detoxification mechanisms resulting from imidacloprid and chlorothalonil exposure.

The Colorado potato beetle, Leptinotarsa decemlineata (Say), is an agricultural pest of commercial potatoes in parts of North America, Europe, and Asia. Plant protection strategies within this geographic range employ a variety of pesticides to combat not only the insect, but also plant pathogens. Previous research has shown that field populations of Leptinotarsa decemlineata have a chronological history of resistance development to a suite of insecticides, including the Group 4A neonicotinoids. The aim of this study is to contextualize the transcriptomic response of Leptinotarsa decemlineata when exposed to the neonicotinoid insecticide imidacloprid, or the fungicides boscalid or chlorothalonil, in order to determine whether these compounds induce similar detoxification mechanisms. We found that chlorothalonil and imidacloprid induced similar patterns of transcript expression, including the up-regulation of a cytochrome p450 and a UDP-glucuronosyltransferase transcript, which belong to protein families associated with xenobiotic metabolism. Further, transcriptomic responses varied among individuals within the same treatment group, suggesting individual insects' responses vary within a population and may cope with chemical stressors in a variety of manners.

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

The Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say), is an agricultural pest of solanaceous crops which has developed insecticide resistance at an alarming rate. Up to this point, little consideration has been given to unintended, or inadvertent effects that non-insecticide xenobiotics may have on insecticide susceptibility in L. decemlineata. Fungicides, such as chlorothalonil and boscalid, are often used to control fungal pathogens in potato fields and are applied at regular intervals when L. decemlineata populations are present in the crop. In order to determine whether fungicide use may be associated with elevated levels of insecticide resistance in L. decemlineata, we examined phenotypic responses in L. decemlineata to the fungicides chlorothalonil and boscalid. Using enzymatic and transcript abundance investigations, we also examined modes of molecular detoxification in response to both insecticide (imidacloprid) and fungicide (boscalid and chlorothalonil) application to more specifically determine if fungicides and insecticides induce similar metabolic detoxification mechanisms. Both chlorothalonil and boscalid exposure induced a phenotypic, enzymatic and transcript response in L. decemlineata which correlates with known mechanisms of insecticide resistance.

Leafhopper-Induced Activation of the Jasmonic Acid Response Benefits Salmonella enterica in a Flagellum-Dependent Manner.

Enteric human pathogens such as Salmonella enterica are typically studied in the context of their animal hosts, but it has become apparent that these bacteria spend a significant portion of their life cycle on plants. S. enterica survives the numerous stresses common to a plant niche, including defense responses, water and nutrient limitation, and exposure to UV irradiation leading to an increased potential for human disease. In fact, S. enterica is estimated to cause over one million cases of foodborne illness each year in the United States with 20% of those cases resulting from consumption of contaminated produce. Although S. enterica successfully persists in the plant environment, phytobacterial infection by Pectobacterium carotovorum or Xanthomonas spp. increases S. enterica survival and infrequently leads to growth on infected plants. The co-association of phytophagous insects, such as the Aster leafhopper, Macrosteles quadrilineatus, results in S. enterica populations that persist at higher levels for longer periods of time when compared to plants treated with S. enterica alone. We hypothesized that leafhoppers increase S. enterica persistence by altering the plant defense response to the benefit of the bacteria. Leafhopper infestation activated the jasmonic acid (JA) defense response while S. enterica colonization triggered the salicylic acid (SA) response. In tomato plants co-treated with S. enterica and leafhoppers, both JA- and SA-inducible genes were activated, suggesting that the presence of leafhoppers may affect the crosstalk that occurs between the two immune response pathways. To rule out the possibility that leafhoppers provide additional benefits to S. enterica, plants were treated with a chemical JA analog to activate the immune response in the absence of leafhoppers. Although bacterial populations continue to decline over time, analog treatment significantly increased bacterial persistence on the leaf surface. Bacterial mutant analysis determined that the bacterial flagellum, whether functional or not, was required for increased S. enterica survival after analog treatment. By investigating the interaction between this human pathogen, a common phytophagous insect, and their plant host, we hope to elucidate the mechanisms promoting S. enterica survival on plants and provide information to be used in the development of new food safety intervention strategies.

Landscape genomics of Colorado potato beetle provides evidence of polygenic adaptation to insecticides.

The ability of insect pests to rapidly and repeatedly adapt to insecticides has long challenged entomologists and evolutionary biologists. Since Crow's seminal paper on insecticide resistance in 1957, new data and insights continue to emerge that are relevant to the old questions about how insecticide resistance evolves: such as whether it is predominantly mono- or polygenic, and evolving from standing vs. de novo genetic variation. Many studies support the monogenic hypothesis, and current management recommendations assume single- or two-locus models. But inferences could be improved by integrating data from a broader sample of pest populations and genomes. Here, we generate evidence relevant to these questions by applying a landscape genomics framework to the study of insecticide resistance in a major agricultural pest, Colorado potato beetle, Leptinotarsa decemlineata (Say). Genome-environment association tests using genomic variation from 16 populations spanning gradients of landscape variables associated with insecticide exposure over time revealed 42 strong candidate insecticide resistance genes, with potentially overlapping roles in multiple resistance mechanisms. Measurements of resistance to a widely used insecticide, imidacloprid, among 47 L. decemlineata populations revealed heterogeneity at a small (2 km) scale and no spatial signature of origin or spread throughout the landscape. Analysis of nucleotide diversity suggested candidate resistance loci have undergone varying degrees of selective sweeps, often maintaining similar levels of nucleotide diversity to neutral loci. This study suggests that many genes are involved in insecticide resistance in L. decemlineata and that resistance likely evolves from both de novo and standing genetic variation.

RNA interference of three up-regulated transcripts associated with insecticide resistance in an imidacloprid resistant population of Leptinotarsa decemlineata.

The Colorado potato beetle, Leptinotarsa decemlineata (Say), is a major agricultural pest of potatoes in the Central Sands production region of Wisconsin. Previous studies have shown that populations of L. decemlineata have become resistant to many classes of insecticides, including the neonicotinoid insecticide, imidacloprid. Furthermore, L. decemlineata has multiple mechanisms of resistance to deal with a pesticide insult, including enhanced metabolic detoxification by cytochrome p450s and glutathione S-transferases. With recent advances in the transcriptomic analysis of imidacloprid susceptible and resistant L. decemlineata populations, it is possible to investigate the role of candidate genes involved in imidacloprid resistance. A recently annotated transcriptome analysis of L. decemlineata was obtained from select populations of L. decemlineata collected in the Central Sands potato production region, which revealed a subset of mRNA transcripts constitutively up-regulated in resistant populations. We hypothesize that a portion of the up-regulated transcripts encoding for genes within the resistant populations also encode for pesticide resistance and can be suppressed to re-establish a susceptible phenotype. In this study, a discrete set of three up-regulated targets were selected for RNA interference experiments using a resistant L. decemlineata population. Following the successful suppression of transcripts encoding for a cytochrome p450, a cuticular protein, and a glutathione synthetase protein in a select L. decemlineata population, we observed reductions in measured resistance to imidacloprid that strongly suggest these genes control essential steps in imidacloprid metabolism in these field populations.

Seasonal Population Dynamics of Thrips (Thysanoptera) in Wisconsin and Iowa Soybean Fields.

With the discovery of Neohydatothrips variabilis (Beach) as a vector of Soybean vein necrosis virus (Family Bunyaviridae Genus Tospovirus), a relatively new pathogen of soybean, a multiyear study was initiated in Wisconsin (2013 and 2014) and Iowa (2014 and 2015) to determine the phenology and species composition of thrips in soybean fields. Yellow sticky card traps were used to sample thrips at regular intervals in five counties within each state's primary soybean-growing region. The assemblage of species present in Wisconsin was determined in all site-years, revealing that N. variabilis and other known vectors of tospoviruses were a relatively small percentage of the total thrips captures in 2013 (1.6%) and 2014 (3.6%). A repeated measures analysis was conducted on cumulative proportion thrips capture data within each state's sampling year to investigate differences in phenology, and standardized cumulative insect days were analyzed between sampling years within each state to determine differences in the relative magnitude of populations. Distinct seasonal trends were not detected based on location, as originally hypothesized, and thrips populations varied significantly among locations and between years. These results suggest that thrips populations may be overwintering in northern climates instead of relying solely on migrations to colonize northern soybean fields.