A draft Diabrotica virgifera virgifera genome: insights into control and host plant adaption by a major maize pest insect.

BACKGROUND: Adaptations by arthropod pests to host plant defenses of crops determine their impacts on agricultural production. The larval host range of western corn rootworm, Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae), is restricted to maize and a few grasses. Resistance of D. v. virgifera to crop rotation practices and multiple insecticides contributes to its status as the most damaging pest of cultivated maize in North America and Europe. The extent to which adaptations by this pest contributes to host plant specialization remains unknown. RESULTS: A 2.42 Gb draft D. v. virgifera genome, Dvir_v2.0, was assembled from short shotgun reads and scaffolded using long-insert mate-pair, transcriptome and linked read data. K-mer analysis predicted a repeat content of ≥ 61.5%. Ortholog assignments for Dvir_2.0 RefSeq models predict a greater number of species-specific gene duplications, including expansions in ATP binding cassette transporter and chemosensory gene families, than in other Coleoptera. A majority of annotated D. v. virgifera cytochrome P450s belong to CYP4, 6, and 9 clades. A total of 5,404 transcripts were differentially-expressed between D. v. virgifera larvae fed maize roots compared to alternative host (Miscanthus), a marginal host (Panicum virgatum), a poor host (Sorghum bicolor) and starvation treatments; Among differentially-expressed transcripts, 1,908 were shared across treatments and the least number were between Miscanthus compared to maize. Differentially-expressed transcripts were enriched for putative spliceosome, proteosome, and intracellular transport functions. General stress pathway functions were unique and enriched among up-regulated transcripts in marginal host, poor host, and starvation responses compared to responses on primary (maize) and alternate hosts. CONCLUSIONS: Manual annotation of D. v. virgifera Dvir_2.0 RefSeq models predicted expansion of paralogs with gene families putatively involved in insecticide resistance and chemosensory perception. Our study also suggests that adaptations of D. v. virgifera larvae to feeding on an alternate host plant invoke fewer transcriptional changes compared to marginal or poor hosts. The shared up-regulation of stress response pathways between marginal host and poor host, and starvation treatments may reflect nutrient deprivation. This study provides insight into transcriptomic responses of larval feeding on different host plants and resources for genomic research on this economically significant pest of maize.

Monarch Butterfly Ecology, Behavior, and Vulnerabilities in North Central United States Agricultural Landscapes.

The North American monarch butterfly (Danaus plexippus) is a candidate species for listing under the Endangered Species Act. Multiple factors are associated with the decline in the eastern population, including the loss of breeding and foraging habitat and pesticide use. Establishing habitat in agricultural landscapes of the North Central region of the United States is critical to increasing reproduction during the summer. We integrated spatially explicit modeling with empirical movement ecology and pesticide toxicology studies to simulate population outcomes for different habitat establishment scenarios. Because of their mobility, we conclude that breeding monarchs in the North Central states should be resilient to pesticide use and habitat fragmentation. Consequently, we predict that adult monarch recruitment can be enhanced even if new habitat is established near pesticide-treated crop fields. Our research has improved the understanding of monarch population dynamics at the landscape scale by examining the interactions among monarch movement ecology, habitat fragmentation, and pesticide use.

Up-regulation of apoptotic- and cell survival-related gene pathways following exposures of western corn rootworm to B. thuringiensis crystalline pesticidal proteins in transgenic maize roots.

BACKGROUND: Resistance of pest insect species to insecticides, including B. thuringiensis (Bt) pesticidal proteins expressed by transgenic plants, is a threat to global food security. Despite the western corn rootworm, Diabrotica virgifera virgifera, being a major pest of maize and having populations showing increasing levels of resistance to hybrids expressing Bt pesticidal proteins, the cell mechanisms leading to mortality are not fully understood. RESULTS: Twenty unique RNA-seq libraries from the Bt susceptible D. v. virgifera inbred line Ped12, representing all growth stages and a range of different adult and larval exposures, were assembled into a reference transcriptome. Ten-day exposures of Ped12 larvae to transgenic Bt Cry3Bb1 and Gpp34/Tpp35Ab1 maize roots showed significant differential expression of 1055 and 1374 transcripts, respectively, compared to cohorts on non-Bt maize. Among these, 696 were differentially expressed in both Cry3Bb1 and Gpp34/Tpp35Ab1 maize exposures. Differentially-expressed transcripts encoded protein domains putatively involved in detoxification, metabolism, binding, and transport, were, in part, shared among transcripts that changed significantly following exposures to the entomopathogens Heterorhabditis bacteriophora and Metarhizium anisopliae. Differentially expressed transcripts in common between Bt and entomopathogen treatments encode proteins in general stress response pathways, including putative Bt binding receptors from the ATP binding cassette transporter superfamily. Putative caspases, pro- and anti-apoptotic factors, as well as endoplasmic reticulum (ER) stress-response factors were identified among transcripts uniquely up-regulated following exposure to either Bt protein. CONCLUSIONS: Our study suggests that the up-regulation of genes involved in ER stress management and apoptotic progression may be important in determining cell fate following exposure of susceptible D. v. virgifera larvae to Bt maize roots. This study provides novel insights into insect response to Bt intoxication, and a possible framework for future investigations of resistance mechanisms.

Influence of host plant, geography and pheromone strain on genomic differentiation in sympatric populations of Ostrinia nubilalis.

Patterns of mating for the European corn borer (Ostrinia nubilalis) moth depend in part on variation in sex-pheromone blend. The ratio of (E)-11- and (Z)-11-tetradecenyl acetate (E11- and Z11-14:OAc) in the pheromone blend that females produce and males respond to differs between strains of O. nubilalis. Populations also vary in female oviposition preference for and larval performance on maize (C4) and nonmaize (C3) host plants. The relative contributions of sexual and ecological trait variation to the genetic structure of O. nubilalis remains unknown. Host-plant use (13 C/14 C ratios) and genetic differentiation were estimated among sympatric E and Z pheromone strain O. nubilalis males collected in sex-pheromone baited traps at 12 locations in Pennsylvania and New York between 2007 and 2010. Among genotypes at 65 single nucleotide polymorphism marker loci, variance at a position in the pheromone gland fatty acyl-reductase (pgfar) gene at the locus responsible for determining female pheromone ratio (Pher) explained 64% of the total genetic differentiation between males attracted to different pheromones (male response, Resp), providing evidence of sexual inter-selection at these unlinked loci. Principal coordinate, Bayesian clustering, and distance-based redundancy analysis (dbRDA) demonstrate that host plant history or geography does not significantly contribute to population variation or differentiation among males. In contrast, these analyses indicate that pheromone response and pgfar-defined strain contribute significantly to population genetic differentiation. This study suggests that behavioural divergence probably plays a larger role in driving genetic variation compared to host plant-defined ecological adaptation.

Early Detection and Mitigation of Resistance to Bt Maize by Western Corn Rootworm (Coleoptera: Chrysomelidae).

Transgenic Bt maize that produces less than a high-dose has been widely adopted and presents considerable insect resistance management (IRM) challenges. Western corn rootworm, Diabrotica virgifera virgifera LeConte, has rapidly evolved resistance to Bt maize in the field, leading to local loss of efficacy for some corn rootworm Bt maize events. Documenting and responding to this resistance has been complicated by a lack of rapid diagnostic bioassays and by regulatory triggers that hinder timely and effective management responses. These failures are of great concern to the scientific and agricultural community. Specific challenges posed by western corn rootworm resistance to Bt maize, and more general concerns around Bt crops that produce less than a high-dose of Bt toxin, have caused uncertainty around current IRM protocols. More than 15 years of experience with IRM has shown that high-dose and refuge-based IRM is not applicable to Bt crops that produce less than a high-dose. Adaptive IRM approaches and pro-active, integrated IRM-pest management strategies are needed and should be in place before release of new technologies that produce less than a high-dose. We suggest changes in IRM strategies to preserve the utility of corn rootworm Bt maize by 1) targeting local resistance management earlier in the sequence of responses to resistance and 2) developing area-wide criteria to address widespread economic losses. We also favor consideration of policies and programs to counteract economic forces that are contributing to rapid resistance evolution.

Haplotype Profile Comparisons Between Spodoptera frugiperda (Lepidoptera: Noctuidae) Populations From Mexico With Those From Puerto Rico, South America, and the United States and Their Implications to Migratory Behavior.

Fall armyworm [Spodoptera frugiperda (J. E. Smith)] is a major economic pest throughout the Western Hemisphere of maize, cotton, sorghum, and a variety of agricultural grasses and vegetable crops. Previous studies demonstrated extensive annual migrations occurring as far north as Canada from overwintering locations in southern Florida and Texas. In contrast, migratory behavior in the rest of the hemisphere is largely uncharacterized. Understanding the migration patterns of fall armyworm will facilitate efforts to predict the spread of pesticide resistance traits that repeatedly arise in this species and assess the consequences of changing climatic trends on the infestation range. Four independent fall armyworm colonies derived from widely separated populations in Mexico and two field collections were examined for their mitochondrial cytochrome oxidase I (COI) gene haplotypes and compared with other locations. The Mexico populations were most similar in their haplotype profile to those from Texas and South America, but also displayed some distinctive features. The data extend the haplotype distribution map in the Western Hemisphere and confirm that the previously observed regional differences in haplotype frequencies are stable over time. The Mexico collections were associated with haplotypes rarely found elsewhere, suggesting limited migratory interactions with foreign populations, including those in neighboring Texas.

Aseasonal, undirected migration in insects: 'Invisible' but common.

Many insect pests are long-distance migrants, moving from lower latitudes where they overwinter to higher latitudes in spring to exploit superabundant, but seasonally ephemeral, host crops. These seasonal long-distance migration events are relatively easy to recognize, and justifiably garner much research attention. Evidence indicates several pest species that overwinter in diapause, and thus inhabit a year-round range, also engage in migratory flight, which is somewhat "invisible" because displacement is nondirectional and terminates among conspecifics. Support for aseasonal, undirected migration is related to recognizing true migratory flight behavior, which differs fundamentally from most other kinds of flight in that it is nonappetitive. Migrating adults are not searching for resources and migratory flight is not arrested by encounters with potential resources. The population-level consequence of aseasonal, undirected migration is spatial mixing of individuals within the larger metapopulation, which has important implications for population dynamics, gene flow, pest management, and insect resistance management.

Critical Facets of European Corn Borer Adult Movement Ecology Relevant to Mitigating Field Resistance to Bt-Corn.

The European corn borer (Ostrinia nubilalis, Hübner) has been managed successfully in North America since 1996 with transgenic Bt-corn. However, field-evolved resistance to all four available insecticidal Bt proteins has been detected in four provinces of Canada since 2018. Evidence suggests resistance may be spreading and evolving independently in scattered hotspots. Evolution and spread of resistance are functions of gene flow, and therefore dispersal, so design of effective resistance management and mitigation plans must take insect movement into account. Recent advances in characterizing European corn borer movement ecology have revealed a number of surprises, chief among them that a large percentage of adults disperse from the natal field via true migratory behavior, most before mating. This undermines a number of common key assumptions about adult behavior, patterns of movement, and gene flow, and stresses the need to reassess how ecological data are interpreted and how movement in models should be parameterized. While many questions remain concerning adult European corn borer movement ecology, the information currently available is coherent enough to construct a generalized framework useful for estimating the spatial scale required to implement possible Bt-resistance prevention, remediation, and mitigation strategies, and to assess their realistic chances of success.

Evaluation of tolerance to Bacillus thuringiensis toxins among laboratory-reared western bean cutworm (Lepidoptera: Noctuidae).

The western bean cutworm, Striacosta albicosta (Smith) (Lepidoptera: Noctuidae), is a destructive insect pest of dry beans and corn within its native range of western Nebraska and eastern Colorado. However, since the initiation of an eastward range expansion of S. albicosta in the late 1990s, economic damage has been observed in the Midwest, and the species has now reached the Atlantic Coast and Quebec. Economic damage to corn occurs by larval feeding on ears, which is not controlled by commercial transgenic hybrids that express Bacillus thuringiensis (Bt) Cry1Ab, but partial control is observed by corn varieties that express Cry1 F toxins. Inadequate protocols for laboratory rearing of S. albicosta have hindered controlled efficacy experimentation in the laboratory and field. We report an S. albicosta rearing methodology used to maintain alaboratory colony for 12 continuous generations. Rearing procedures were adapted for Bt toxin diet-overlay assays, revealing that S. albicosta larvae exposed to Bt toxin for 14 d were insensitive to Cry1Ab concentrations up to 25,000 ng/cm2. In contrast, neonates exposed to Cry1 F toxin at > or = 250 ng/cm2, showed reduced developmental rates, with estimated effective concentrations of EC50 = 1,187.5 ng/cm2 and EC95 = 10,000.5 ng/cm2. The ability to mass produce this pest insect will enhance fundamental research, including evaluation of control tactics and toxin susceptibility.

Movement Ecology of Adult Western Corn Rootworm: Implications for Management.

Movement of adult western corn rootworm, Diabrotica virgifera virgifera LeConte, is of fundamental importance to this species' population dynamics, ecology, evolution, and interactions with its environment, including cultivated cornfields. Realistic parameterization of dispersal components of models is needed to predict rates of range expansion, development, and spread of resistance to control measures and improve pest and resistance management strategies. However, a coherent understanding of western corn rootworm movement ecology has remained elusive because of conflicting evidence for both short- and long-distance lifetime dispersal, a type of dilemma observed in many species called Reid's paradox. Attempts to resolve this paradox using population genetic strategies to estimate rates of gene flow over space likewise imply greater dispersal distances than direct observations of short-range movement suggest, a dilemma called Slatkin's paradox. Based on the wide-array of available evidence, we present a conceptual model of adult western corn rootworm movement ecology under the premise it is a partially migratory species. We propose that rootworm populations consist of two behavioral phenotypes, resident and migrant. Both engage in local, appetitive flights, but only the migrant phenotype also makes non-appetitive migratory flights, resulting in observed patterns of bimodal dispersal distances and resolution of Reid's and Slatkin's paradoxes.

Distribution of genes and repetitive elements in the Diabrotica virgifera virgifera genome estimated using BAC sequencing.

Feeding damage caused by the western corn rootworm, Diabrotica virgifera virgifera, is destructive to corn plants in North America and Europe where control remains challenging due to evolution of resistance to chemical and transgenic toxins. A BAC library, DvvBAC1, containing 109,486 clones with 104 ± 34.5 kb inserts was created, which has an ~4.56X genome coverage based upon a 2.58 Gb (2.80 pg) flow cytometry-estimated haploid genome size. Paired end sequencing of 1037 BAC inserts produced 1.17 Mb of data (~0.05% genome coverage) and indicated ~9.4 and 16.0% of reads encode, respectively, endogenous genes and transposable elements (TEs). Sequencing genes within BAC full inserts demonstrated that TE densities are high within intergenic and intron regions and contribute to the increased gene size. Comparison of homologous genome regions cloned within different BAC clones indicated that TE movement may cause haplotype variation within the inbred strain. The data presented here indicate that the D. virgifera virgifera genome is large in size and contains a high proportion of repetitive sequence. These BAC sequencing methods that are applicable for characterization of genomes prior to sequencing may likely be valuable resources for genome annotation as well as scaffolding.

Sequences Encoding a Novel Toursvirus Identified from Southern and Northern Corn Rootworms (Coleoptera: Chrysomelidae).

Sequences derived from a novel toursvirus were identified from pooled genomic short read data from U.S. populations of southern corn rootworm (SCR, Diabrotica undecimpunctata howardi Barber) and northern corn rootworm (NCR, Diabrotica barberi Smith & Lawrence). Most viral sequences were identified from the SCR genomic dataset. As proteins encoded by toursvirus sequences from SCR and NCR were almost identical, the contig sets from SCR and NCR were combined to generate 26 contigs. A total of 108,176 bp were assembled from these contigs, with 120 putative toursviral ORFs identified indicating that most of the viral genome had been recovered. These ORFs included all 40 genes that are common to members of the Ascoviridae. Two genes typically present in Ascoviridae (ATP binding cassette transport system permeases and Baculovirus repeated open reading frame), were not detected. There was evidence for transposon insertion in viral sequences at different sites in the two host species. Phylogenetic analyses based on a concatenated set of 45 translated protein sequences clustered toursviruses into a distinct clade. Based on the combined evidence, we propose taxonomic separation of toursviruses from Ascoviridae.

Patterns of Microbiome Composition Vary Across Spatial Scales in a Specialist Insect.

Microbial communities associated with animals vary based on both intrinsic and extrinsic factors. Of many possible determinants affecting microbiome composition, host phylogeny, host diet, and local environment are the most important. How these factors interact across spatial scales is not well understood. Here, we seek to identify the main influences on microbiome composition in a specialist insect, the western corn rootworm (WCR; Diabrotica virgifera virgifera), by analyzing the bacterial communities of adults collected from their obligate host plant, corn (Zea mays), across several geographic locations and comparing the patterns in communities to its congeneric species, the northern corn rootworm (NCR; Diabrotica barberi). We found that bacterial communities of WCR and NCR shared a portion of their bacterial communities even when collected from disparate locations. However, within each species, the location of collection significantly influenced the composition of their microbiome. Correlations of geographic distance between sites with WCR bacterial community composition revealed different patterns at different spatial scales. Community similarity decreased with increased geographic distance at smaller spatial scales (~25 km between the nearest sites). At broad spatial scales (>200 km), community composition was not correlated with distances between sites, but instead reflected the historical invasion path of WCR across the United States. These results suggest bacterial communities are structured directly by dispersal dynamics at small, regional spatial scales, while landscape-level genetic or environmental differences may drive community composition across broad spatial scales in this specialist insect.

Population Development, Fecundity, and Flight of Spodoptera frugiperda (Lepidoptera: Noctuidae) Reared on Three Green Manure Crops: Implications For an Ecologically Based Pest Management Approach in China.

The fall armyworm, Spodoptera frugiperda (Smith), is an invasive pest of cereal crops that now inhabits southern China year-round. Cultivation of crops unsuitable as host plants has been an effective pest management strategy for some insect pests, but the effects of green manure crops on S. frugiperda have not been investigated. An age-stage two-sex life table and tethered flight performance of S. frugiperda reared on different green manure species were obtained, and a population dynamics model established. Developmental durations of stages, survival rates, and fecundities of S. frugiperda differed significantly depending on host plant. Larvae fed Astragalus sinicus L. did not complete development. Although some larvae fed Vicia villosa Roth and Vicia sativa L. completed development, generation time was significantly prolonged, egg production was halved, and net reproductive rate decreased to 31% and 3% of those reared on corn, respectively. Survival rates of early-instars fed V. villosa and V. sativa were significantly lower than those fed corn. Population dynamics projections over 90 d showed the number of generations of S. frugiperda fed on V. villosa and V. sativa was reduced compared to those reared on corn. Flight performance of S. frugiperda reared on V. villosa decreased significantly compared to corn. Our results show that the three green manure species are unsuitable host plants for S. frugiperda. Therefore, reduction of corn production in southern China through rotation with these green manure crops may be a feasible method of ecological management of this major corn pest in China.

The oriental armyworm genome yields insights into the long-distance migration of noctuid moths.

The oriental armyworm, Mythimna separata, is known for its long-distance seasonal migration and environment-dependent phase polymorphisms. Here, we present a chromosome-level genome reference and integrate multi-omics, functional genetics, and behavioral assays to explore the genetic bases of the hallmark traits of M. separata migration. Gene family comparisons show expansion of gustatory receptor genes in this cereal crop pest. Functional investigation of magnetoreception-related genes and associated flight behaviors suggest that M. separata may use the geomagnetic field to guide orientation in its nocturnal flight. Comparative transcriptome characterizes a suite of genes that may confer the observed plasticity between phases, including genes involved in protein processing, hormone regulation, and dopamine metabolism. We further report molecular signatures that underlie the dynamic regulation of a migratory syndrome coordinating reproduction and flight. Our study yields insights into environment-dependent developmental plasticity in moths and advances our understanding of long-distance migration in nocturnal insect pests.

Increased survival of western corn rootworm on transgenic corn within three generations of on-plant greenhouse selection.

To delay evolution of insect resistance to transgenic crops producing Bacillus thuringiensis (Bt) toxins, nearby "refuges" of host plants not producing Bt toxins are required in many regions. Such refuges are expected to be most effective in slowing resistance when the toxin concentration in Bt crops is high enough to kill all or nearly all insects heterozygous for resistance. However, Bt corn, Zea mays, introduced recently does not meet this "high-dose" criterion for control of western corn rootworm (WCR), Diabrotica virgifera virgifera. A greenhouse method of rearing WCR on transgenic corn expressing the Cry3Bb1 protein was used in which approximately 25% of previously unexposed larvae survived relative to isoline survival (compared to 1-4% in the field). After three generations of full larval rearing on Bt corn (Constant-exposure colony), WCR larval survival was equivalent on Bt corn and isoline corn in greenhouse trials, and the LC(50) was 22-fold greater for the Constant-exposure colony than for the Control colony in diet bioassays with Cry3Bb1 protein on artificial diet. After six generations of greenhouse selection, the ratio of larval recovery on Bt corn to isoline corn in the field was 11.7-fold greater for the Constant-exposure colony than the Control colony. Removal from selection for six generations did not decrease survival on Bt corn in the greenhouse. The results suggest that rapid response to selection is possible in the absence of mating with unexposed beetles, emphasizing the importance of effective refuges for resistance management.

Starvation on First or Second Day of Adulthood Reverses Larval-Stage Decision to Migrate in Beet Webworm (Lepidoptera: Pyralidae).

A facultative commitment to adult migration in the larval stage can be modified again after adult emergence in some Lepidoptera when influenced by an appropriate environmental cue during a sensitive stage. This phenomenon is termed secondary regulation of migration. The sensitive stage in adult beet webworm, Loxostege sticticalis L. (Lepidoptera: Pyralidae), was determined experimentally by starvation of presumed migrant females reared from gregarious-phase larvae (induced by crowding at 10 larvae per 650-ml jar). When presumed migrant adults were starved for 24 h on either of the first 2 d after emergence, the preoviposition period was shortened. In contrast, preoviposition periods were not significantly shortened for migrants starved on day 3 or when starvation lasted for more than 1 d after emergence. Because the preoviposition period corresponds to the migratory period in beet webworm, the results suggest that the first 2 d of adult life in the beet webworm is the sensitive stage during which presumed migrants can be switched to residents by an appropriate environmental cue. During the sensitive stage or not, starvation did not influence lifetime fecundity, oviposition period, longevity, or hatching rate of eggs laid by the starvation-stressed moths. Starvation on the first day also increased tethered flight performance and accelerated both flight muscle and ovarian development. The results suggest that a pulse of starvation in the sensitive period may inhibit the expected migration by accelerating and compressing the cycle of migratory flight muscle development and degeneration, while accelerating ovarian development, which is normally suppressed until after migration.

Nudivirus Sequences Identified from the Southern and Western Corn Rootworms (Coleoptera: Chrysomelidae).

Analysis of pooled genomic short read sequence data revealed the presence of nudivirus-derived sequences from U.S. populations of both southern corn rootworm (SCR, Diabrotica undecimpunctata howardi Barber) and western corn rootworm (WCR, Diabrotica virgifera virgifera LeConte). A near complete nudivirus genome sequence was assembled from sequence data for an SCR population with relatively high viral titers. A total of 147,179 bp was assembled from five contigs that collectively encode 109 putative open reading frames (ORFs) including 20 nudivirus core genes. In contrast, genome sequence recovery was incomplete for a second nudivirus from WCR, although sequences derived from this virus were present in three geographically dispersed populations. Only 48,989 bp were assembled with 48 putative ORFs including 13 core genes, representing about 20% of a typical nudivirus genome. Phylogenetic analysis indicated that both corn rootworm nudiviruses grouped with the third known nudivirus of beetles, Oryctes rhinoceros nudivirus in the genus Alphanudivirus. On the basis of phylogenetic and additional analyses, we propose further taxonomic separation of nudiviruses within Alphanudivirus and Betanudivirus into two subfamilies and five genera. Identification of nudivirus-derived sequences from two species of corn rootworm highlights the diversity of viruses associated with these agricultural insect pests.

Oviposition Response of Monarch Butterfly (Lepidoptera: Nymphalidae) to Imidacloprid-Treated Milkweed.

Monarch butterfly (Danaus plexippus) populations have declined over the last two decades, attributable in part to declines in its larval host plant, milkweed (Asclepias spp.), across its breeding range. Conservation efforts in the United States call for restoration of 1.3 billion milkweed stems into the Midwestern landscape. Reaching this goal will require habitat establishment in marginal croplands, where there is a high potential for exposure to agrochemicals. Corn and soybean crops may be treated with neonicotinoid insecticides systemically or through foliar applications to provide protection against insect pests. Here, we investigate whether ovipositing monarchs discriminate against milkweed plants exposed to the neonicotinoid insecticide imidacloprid, either systemically or through foliar application. In our first experiment, we placed gravid females in enclosures containing a choice of two cut stems for oviposition: one in 15 ml of a 0.5 mg/ml aqueous solution of imidacloprid and one in 15 ml water. In a second experiment, females were given a choice of milkweed plants whose leaves were treated with 30 µl of a 0.825 mg/ml imidacloprid-surfactant solution or plants treated with surfactant alone. To evaluate oviposition preference, we counted and removed eggs from all plants daily for 3 d. We also collected video data on a subset of butterflies to evaluate landing behavior. Results indicate that neither systemic nor foliar treatment with imidacloprid influenced oviposition behavior in female monarchs. The implications of these findings for monarch conservation practices will be informed by the results of ongoing egg and larval toxicity studies.

Key factors involved in reduction of damage to sunflower by the European sunflower moth in China through late planting.

The European sunflower moth, Homoesoma nebulellum (Denis et Schiffermüller), emerged as a major new pest in Bayannur, China, in 2006. Insecticidal control with a single application is problematic because timing is critical, and multiple applications increase production and environmental costs. Management of H. nebulellum by planting date adjustment can be effective, but the optimal time window for late planting is unknown. Natural levels of H. nebulellum infestation were compared among sunflowers planted on five dates from April 25 to June 5 in two years, and the relationship between timing of adult abundance and flowering assessed. Delaying planting of sunflower from the traditional planting period of April 25 -May 5 to May 15 -June 5 significantly decreased damage by H. nebulellum. Seed infestation rate was 30-40 times higher, and number of larvae/head 75-100 times higher in the earliest two plantings than in the latest two. Within two years of implementing delayed planting in Bayannur city, infestation area decreased from 72% in 2006 to 1.5% in 2008, and production losses decreased from 4.5 ton/ha in 2006 to 0.36 ton/ha in 2008, a 97% decrease compared to 2006. Moreover, the infestation area caused by H. nebulellum was continuously controlled below 5.3% of the planting area since 2008. We found the overlap between the first two days of flowering and peak adult presence was the key factor influencing level of damage caused by H. nebulellum. Because the number of eggs laid in the first two days of flowering accounted for 68% of the total, and sunflower seed infestation rate was positively correlated with the number of trapped adults weighted by proportion of daily oviposition. Oviposition of the majority of eggs in the first two days of flowering suggests an evolutionary mechanism whereby females choose host plants most conducive to larval development, consistent with the preference-performance hypothesis.