Nutritional resources used by the invasive maize pest Diabrotica virgifera virgifera in its new South-east-European distribution range

Food utilization by adults of the invasive maize (Zea mays L.) (Poaceae) pest western corn rootworm (WCR; Diabrotica virgifera virgifera LeConte) (Coleoptera: Chrysomelidae) was investigated in the south‐eastern part of its new European distribution range. At weekly intervals over a 10‐week period, 10 beetles per field were collected from six fields that had a high abundance of flowering weeds and six fields with a low abundance of flowering weeds, with the aim of understanding adult feeding behaviour in Europe and comparing this behaviour with North American WCR. Gut content analysis was performed to determine the use of maize tissue and weed pollen with regard to maize phenology. Furthermore, all pollen found within the gut was quantified and identified to plant species level. The use of maize tissue by adult WCR changed over time according to maize phenology. Pollen originating from plants other than maize was used more frequently as the maize matured. Adults fed on pollen from 19 of 25 different plant species found in maize fields and showed a preference for the pollen of specific weeds. Pollen from weed species was found more often in beetles from fields with a high abundance of weeds compared to beetles from fields with a low abundance of weeds. Females consumed greater amounts of pollen than males, the latter feeding on a wider diversity of host plants. The pollen resources used by adult WCR in Hungary were more diverse compared to WCR in the USA, which may contribute to the invasion success of WCR in Europe.     

Evidence of Field-Evolved Resistance to Bifenthrin in Western Corn Rootworm (Diabrotica virgifera virgifera LeConte) Populations in Western Nebraska and Kansas

Pyrethroid insecticides have been used to control larvae or adults of the western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, a key pest of field corn in the United States. In response to reports of reduced efficacy of pyrethroids in WCR management programs in southwestern areas of Nebraska and Kansas the present research was designed to establish a baseline of susceptibility to the pyrethroid insecticide, bifenthrin, using susceptible laboratory populations and to compare this baseline with susceptibility of field populations. Concentration-response bioassays were performed to estimate the baseline susceptibility. From the baseline data, a diagnostic concentration (LC₉₉) was determined and used to test adults of both laboratory and field populations. Larval susceptibility was also tested using both laboratory and field populations. Significant differences were recorded in adult and larval susceptibility among WCR field and laboratory populations. The highest LC₅₀ for WCR adults was observed in populations from Keith 2 and Chase Counties, NE, with LC₅₀s of 2.2 and 1.38 μg/vial, respectively, and Finney County 1, KS, with 1.43 μg/vial, as compared to a laboratory non-diapause population (0.24 μg/vial). For larvae, significant differences between WCR field and laboratory populations were also recorded. Significant differences in mortalities at the diagnostic bifenthrin concentration (LC₉₉) were observed among WCR adult populations with western Corn Belt populations exhibiting lower susceptibility to bifenthrin, especially in southwestern Nebraska and southwestern Kansas. This study provides evidence that resistance to bifenthrin is evolving in field populations that have been exposed for multiple years to pyrethroid insecticides. Implications to sustainable rootworm management are discussed.     

Quantifying rates of random mating in western corn rootworm emerging from Cry3Bb1‐expressing and refuge maize in field cages

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is the most significant pest of field maize, Zea mays L. (Poaceae), in the USA. Maize plants expressing Bt toxins targeting the corn rootworm complex have been widely adopted and are the primary insecticidal control measure for this pest in North America. Insect resistance management tactics using various refuge structures have been adopted to ensure Bt products will retain durability. An assumption of the refuge strategy is that males and females emerging from Bt and refuge plantings mate randomly; this has not been tested in the field. We conducted cage studies using field populations of WCR in Indiana, USA, to generate empirical field data on mating rates between beetles emerging from Cry3Bb1‐expressing Bt and refuge maize plants. Two refuge configurations were tested; all refuge plants were labeled using the stable isotope ¹⁵N. This mark persists in adult beetles after eclosion, allowing for collection and analysis of isotopic ratios of all beetles. Additional data collected included adult emergence rates, timing and sex ratios for each of the treatments, and head capsule size and dry weights of beetles collected. Treatment had a significant effect on dry weight; mean dry weight decreased in Bt‐only treatments. Fisher's exact test of proportions of mating pairs of refuge and Bt insects indicated that mating was not random in 20% strip refuges and 5% seed blend treatments. We found high percentages of beetles that fed on Bt‐expressing plants as larvae, suggesting that mating between resistant beetles may not be rare even if random mating did occur.     

Behaviour and ecology of the western corn rootworm (Diabrotica virgifera virgifera LeConte)

1 The western corn rootworm (WCR) is a historic pest with a legacy of resistance and behavioural plasticity. Its behaviour and nutritional ecology are important to rootworm management. The success of the most effective and environmentally benign rootworm management method, annual crop rotation, was based on an understanding of rootworm behaviour and host–plant relationships. Enthusiastic adoption of crop rotation, provided excellent rootworm management, but also selected for behavioural resistance to this cultural control.
2 Though well-studied, significant gaps in WCR biology remain. Understanding the topics reviewed here (mating behaviour, nutritional ecology, larval and adult movement, oviposition, alternate host use, and chemical ecology) is a starting point for adapting integrated pest management and insect resistance management (IRM) to an expanding WCR threat. A presentation of significant questions and areas in need of further study follow each topic.
3 The expansion of WCR populations into Europe exposes this pest to new environmental and regulatory conditions that may influence its behaviour and ecology. Reviewing the state of current knowledge provides a starting point of reference for researchers and pest management decision-makers in North America and Europe.
4 The trend toward increasing adoption of transgenic maize will place an increasing premium on understanding WCR behaviour. IRM plans designed to promote sustainable deployment of transgenic hybrids are grounded on assumptions about WCR movement, mating and ovipositional behaviour. Preserving the utility of new and old management options will continue to depend on a thorough understanding of WCR biology, even as the ecological circumstances and geography of WCR problems become more complex.     

A selective insecticidal protein from Pseudomonas mosselii for corn rootworm control

The coleopteran insect western corn rootworm (WCR, Diabrotica virgifera virgifera) is an economically important pest in North America and Europe. Transgenic corn plants producing Bacillus thuringiensis (Bt) insecticidal proteins have been useful against this devastating pest, but evolution of resistance has reduced their efficacy. Here, we report the discovery of a novel insecticidal protein, PIP‐47Aa, from an isolate of Pseudomonas mosselii. PIP‐47Aa sequence shows no shared motifs, domains or signatures with other known proteins. Recombinant PIP‐47Aa kills WCR, two other corn rootworm pests (Diabrotica barberi and Diabrotica undecimpunctata howardi) and two other beetle species (Diabrotica speciosa and Phyllotreta cruciferae), but it was not toxic to the spotted lady beetle (Coleomegilla maculata) or seven species of Lepidoptera and Hemiptera. Transgenic corn plants expressing PIP‐47Aa show significant protection from root damage by WCR. PIP‐47Aa kills a WCR strain resistant to mCry3A and does not share rootworm midgut binding sites with mCry3A or AfIP‐1A/1B from Alcaligenes that acts like Cry34Ab1/Cry35Ab1. Our results indicate that PIP‐47Aa is a novel insecticidal protein for controlling the corn rootworm pests.     

Establishing an In Vivo Assay System to Identify Components Involved in Environmental RNA Interference in the Western Corn Rootworm

The discovery of environmental RNA interference (RNAi), in which gene expression is suppressed via feeding with double-stranded RNA (dsRNA) molecules, opened the door to the practical application of RNAi-based techniques in crop pest management. The western corn rootworm (WCR, Diabrotica virgifera virgifera) is one of the most devastating corn pests in North America. Interestingly, WCR displays a robust environmental RNAi response, raising the possibility of applying an RNAi-based pest management strategy to this pest. Understanding the molecular mechanisms involved in the WCR environmental RNAi process will allow for determining the rate limiting steps involved with dsRNA toxicity and potential dsRNA resistance mechanisms in WCR. In this study, we have established a two-step in vivo assay system, which allows us to evaluate the involvement of genes in environmental RNAi in WCR. We show that laccase 2 and ebony, critical cuticle pigmentation/tanning genes, can be used as marker genes in our assay system, with ebony being a more stable marker to monitor RNAi activity. In addition, we optimized the dsRNA dose and length for the assay, and confirmed that this assay system is sensitive to detect well-known RNAi components such as Dicer-2 and Argonaute-2. We also evaluated two WCR sid1- like (sil) genes with this assay system. This system will be useful to quickly survey candidate systemic RNAi genes in WCR, and also will be adaptable for a genome-wide RNAi screening to give us an unbiased view of the environmental/systemic RNAi pathway in WCR.     

The ineffectiveness of insecticide seed coatings and planting-time soil insecticides as Diabrotica virgifera virgifera LeConte population suppressors

Abstract:  The effect of any management strategy on pest population levels must be researched and determinations need to be made as to how that strategy might work based on the control objectives. In certain areas of Europe, the objective is to contain or eradicate the western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, population. In order to evaluate the impact of insecticide seed coatings and/or planting-time applications of insecticides as WCR population suppressors, plot trials and large field observations were carried out in Italy over a 5-year period. Larval, pupal and adult densities, along with root damage ratings, were estimated at different locations. Data from these studies revealed that the number of WCR adults emerging from untreated plots did not differ from the number of beetles emerging from those treated with insecticides, whether as seed coating or in-furrow applications. Both seed insecticide coatings (imidacloprid, fipronil, thiamethoxam, tefluthrin) and soil insecticides applied in-furrow (chlorpyrifos, diazinon, tefluthrin) did not reduce the number of beetles emerging from monoculture fields, either in plot trials or large field observations. Observations in the USA had previously shown that soil insecticides applied at planting time partially protected basal roots from economic damage, but did not reduce corn rootworm populations. Similarly, in Europe, it has been demonstrated that not only the application of soil insecticides at planting time but also insecticide seed coatings have no role in the containment and/or eradication of WCR. Although insecticide seed coatings and soil insecticides applied in-furrow may provide protection against economic damage to roots, these management strategies do not have an impact on WCR populations and therefore are useless in WCR containment and eradication programmes.     

Heterorhabditis sp. not attracted to synthetic (E)‐β‐caryophyllene,a volatile emitted by roots upon feeding by corn rootworm

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, is a destructive pest of maize, Zea mays L. in North America and Europe. Larvae and pupae can be controlled with entomopathogenic nematodes (EPN) of the genus Heterorhabditis. When maize is attacked by WCR, the roots of some maize varieties emit (E)‐β‐caryophyllene that attract EPN to the pest larvae. To use synthetic (E)‐β‐caryophyllene in bioassays for the genetic selection of EPN strains with enhanced chemotactic response to a volatile emitted from the damaged root when attacked by larvae of the WCR, different laboratory bioassays were tested. Three sand assays and one agar assay used synthetic (E)‐β‐caryophyllene as an attractant for H. megidis. In none of the assays, attraction of the nematodes to (E)‐β‐caryophyllene was recorded. Possible reasons why (E)‐β‐caryophyllene was not attracting EPN in the bioassays are discussed.     

Resistance evolution to the first generation of genetically modified Diabrotica-active Bt-maize events by western corn rootworm: management and monitoring considerations

Western corn rootworm (Diabrotica virgifera virgifera; WCR) is a major coleopteran maize pest in North America and the EU, and has traditionally been managed through crop rotation and broad-spectrum soil insecticides. Genetically modified Bt-maize offers an additional management tool for WCR and has been valuable in reducing insecticide use and increasing farm income. A concern is that the widespread, repeated, and exclusive deployment of the same Bt-maize transformation event will result in the rapid evolution of resistance in WCR. This publication explores the potential of WCR to evolve resistance to plant-produced Bt-toxins from the first generation of Diabrotica-active Bt-maize events (MON 863 and MON 88017, DAS-59122-7 and MIR604), and whether currently implemented risk management strategies to delay and monitor resistance evolution are appropriate. In twelve of the twelve artificial selection experiments reported, resistant WCR populations were yielded rapidly. Field-selected resistance of WCR to Cry3Bb1 is documented in some US maize growing areas, where an increasing number of cases of unexpected damage of WCR larvae to Bt-maize MON 88017 has been reported. Currently implemented insect resistance management measures for Bt-crops usually rely on the high dose/refuge (HDR) strategy. Evidence (including laboratory, greenhouse and field data) indicates that several conditions contributing to the success of the HDR strategy may not be met for the first generation of Bt-maize events and WCR: (1) the Bt-toxins are expressed heterogeneously at a low-to-moderate dose in roots; (2) resistance alleles may be present at a higher frequency than initially assumed; (3) WCR may mate in a non-random manner; (4) resistance traits could have non-recessive inheritance; and (5) fitness costs may not necessarily be associated with resistance evolution. However, caution must be exercised when extrapolating laboratory and greenhouse results to field conditions. Model predictions suggest that a 20 % refuge of non-Diabrotica-active Bt-maize can delay resistance evolution in WCR under certain conditions. This publication concludes that further research is needed to resolve the remaining scientific uncertainty related to the appropriateness of the HDR in delaying resistance evolution in WCR, resistance monitoring is essential to detect early warning signs indicating resistance evolution in the field, and that integrated pest management reliant on multiple tactics should be deployed to ensure effective long-term corn rootworm management and sustainable use of Bt-maize.     

Use of transgenic plants to measure insect herbivore movement

Use of ingested transgenic corn tissue as a marker for measuring movement of adult Diabrotica virgifera virgifera (LeConte) (Coleoptera: Chrysomelidae; western corn rootworm) was investigated. Laboratory observations of beetles feeding on corn foliage, pollen, silks, or soybean foliage provided background on feeding patterns. The interval between food consumption and its appearance in feces (gut passage time) ranged from 102.7 +/- 11 min for soybean foliage to 56.7 +/- 2.9 min for corn silks. In a laboratory assay, protein expression tests identified the presence of Cry3Bb1 protein inside 50% of adult D. virgifera for up to 16 h after they had last consumed Cry3Bb1 protein-expressing corn silks from 'YieldGard Rootworm' corn plants (Monsanto Co.). Cry3Bb1 protein could not be detected by 32 h postfeeding. The proportion of Cry3Bb1 protein-positive beetles declined linearly with increasing time since feeding on 'YieldGard Rootworm' tissue. Approximately 20% of adult D. virgifera collected near 'YieldGard Rootworm' corn plots tested positive for Cry3Bb1 protein, indicating 'YieldGard Rootworm' tissue consumption within the last 16-32 h. Based on a 16- to 32-h postfeeding detection interval for Cry3Bb1 protein and the distance between 'YieldGard Rootworm' sources and sites where Cry3Bb1-positive insects were collected, 85.3% of males and females moved < or = 4.6-9.1 m/d through R2-R3 stage corn. Among Cry3Bb1-positive adults that left corn and were captured in an adjacent soybean field, 86.4% of males and 93.1% of females moved < or = 4.6-9.1 m/d through soybean. Detection of transgenic plant tissues in mobile insect herbivores is a novel application of biotechnology to the study of insect movement.     

Impact of cysteine proteinase inhibition in midgut fluid and oral secretion on fecundity and pollen consumption of western corn rootworm (Diabrotica virgifera virgifera)

Cysteine proteinases predominate in the midgut fluid (MF) and oral secretion (OS) of adult western corn rootworm (WCR) based on their mild acidic pH optima (pH 6.0), enhanced activities after treatment with thiol reducing agents, and inhibition by selective cysteine proteinase inhibitors (PIs). Four cysteine PIs including E-64, calpeptin, calpain inhibitor II, and leupeptin (also a serine PI) strongly inhibited azocaseinolytic activity in a dose-dependent manner in both the MF and OS. The most significant effect on adult female WCR of cysteine PI consumption with corn pollen was the reduction in fecundity, but female survival was not apparently affected. Mean fresh weights for all PI-fed females were also lower than control groups. All PI-fed groups [E-64, calpain inhibitor I (Cal I) and leupeptin] had a significantly lower daily egg production than respective corn pollen-fed controls. E-64 was more potent than leupeptin and Cal I on inhibiting fecundity, which correlates with their relative anti-proteinase potency in vitro. E-64, Cal I, and leupeptin at 1.5-2 nmol/beetle/day reduced fecundity down to 25-45% of control values. Reduced egg production by PI-fed beetles results from a combination of the direct inhibition of protein digestion and a post-ingestive negative feedback mechanism, which reduces food intake. The supplement of ten essential amino acids into the E-64-treated pollen enhanced up to 3.7-fold the number of eggs laid compared to the E-64-fed group without these amino acids, suggesting that egg production is dependent on the supply of essential amino acids from corn pollen proteolysis.     

Foraging resources of adult Mexican corn rootworm (Coleoptera: Chrysomelidae) in Bell County, Texas

Pollen analyses were used to determine pollen foraging resources of adult Mexican corn rootworms, Diabrotica virgifera zeae Krysan & Smith, captured near Temple, Bell County, TX, in 1996 and 1997. In 1996, adult Mexican corn rootworms were captured in a corn, Zea mays L., field. In 1997, nine locations outside of cornfields were added. Overall, 92% of the beetles (n = 1,323) contained pollen. More than 142,000 pollen grains were counted, representing 45 families, 63 genera, and 27 species. Overall, in 1996, noncorn grass pollen (70%) occurred in the greatest percentage of total pollen followed by corn (17%), then nongrass pollen (13%). In 1997, noncorn grass pollen (76%) had the greatest percentage, then nongrass (18%), and finally corn pollen (6%). Corn pollen was found in 34% of the beetles in 1996 and 26% in 1997. Fifteen Asteraceae taxa were encountered including sunflower, Helianthus annuus L. Thirteen Fabaceae were found including soybean, Glycine max L. Mexican corn rootworm adults foraged mainly on noncorn grass pollen, but also foraged on pollen from a large diversity of plant species, indicating that noncorn pollen may play a role in the food habits of Mexican corn rootworm adults.     

Refuge or Reservoir? The Potential Impacts of the Biofuel Crop Miscanthus x giganteus on a Major Pest of Maize

Background

Interest in the cultivation of biomass crops like the C4 grass Miscanthus x giganteus (Miscanthus) is increasing as global demand for biofuel grows. In the US, Miscanthus is promoted as a crop well-suited to the Corn Belt where it could be cultivated on marginal land interposed with maize and soybean. Interactions (direct and indirect) of Miscanthus, maize, and the major Corn Belt pest of maize, the western corn rootworm, (Diabrotica virgifera virgifera LeConte, WCR) are unknown. Adding a perennial grass/biomass crop to this system is concerning since WCR is adapted to the continuous availability of its grass host, maize (Zea mays).

Methodology/Principal Findings

In a greenhouse and field study, we investigated WCR development and oviposition on Miscanthus. The suitability of Miscanthus for WCR development varied across different WCR populations. Data trends indicate that WCR populations that express behavioural resistance to crop rotation performed as well on Miscanthus as on maize. Over the entire study, total adult WCR emergence from Miscanthus (212 WCR) was 29.6% of that from maize (717 WCR). Adult dry weight was 75–80% that of WCR from maize; female emergence patterns on Miscanthus were similar to females developing on maize. There was no difference in the mean no. of WCR eggs laid at the base of Miscanthus and maize in the field.

Conclusions/Significance

Field oviposition and significant WCR emergence from Miscanthus raises many questions about the nature of likely interactions between Miscanthus, maize and WCR and the potential for Miscanthus to act as a refuge or reservoir for Corn Belt WCR. Responsible consideration of the benefits and risks associated with Corn Belt Miscanthus are critical to protecting an agroecosystem that we depend on for food, feed, and increasingly, fuel. Implications for European agroecosystems in which Miscanthus is being proposed are also discussed in light of the WCR''s recent invasion into Europe.     

Physiological and Cellular Responses Caused by RNAi- Mediated Suppression of Snf7 Orthologue in Western Corn Rootworm (Diabrotica virgifera virgifera) Larvae

Ingestion of double stranded RNA (dsRNA) has been previously demonstrated to be effective in triggering RNA interference (RNAi) in western corn rootworm (WCR, Diabrotica virgifera virgifera LeConte), providing potential novel opportunities for insect pest control. The putative Snf7 homolog of WCR (DvSnf7) has previously been shown to be an effective RNAi target for insect control, as DvSnf7 RNAi leads to lethality of WCR larvae. Snf7 functions as a part of the ESCRT (Endosomal Sorting Complex Required for Transport) pathway which plays a crucial role in cellular housekeeping by internalization, transport, sorting and lysosomal degradation of transmembrane proteins. To understand the effects that lead to death of WCR larvae by DvSnf7 RNAi, we examined some of the distinct cellular processes associated with ESCRT functions such as de-ubiquitination of proteins and autophagy. Our data indicate that ubiquitinated proteins accumulate in DvSnf7 dsRNA-fed larval tissues and that the autophagy process seems to be impaired. These findings suggest that the malfunctioning of these cellular processes in both midgut and fat body tissues triggered by DvSnf7 RNAi were the main effects leading to the death of WCR. This study also illustrates that Snf7 is an essential gene in WCR and its functions are consistent with biological functions described for other eukaryotes.     

Predator responses to novel haemolymph defences of western corn rootworm (Diabrotica virgifera) larvae

Many herbivorous arthropods use defensive chemistry to discourage predators from attacking. This chemistry relies on the ability of predators to rapidly learn to recognize and avoid offensive stimuli. Western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), employs multifaceted chemical defences in its haemolymph, which may contribute significantly to its success as a major economic pest. Here, we test the hypothesis that agrobiont predators can rapidly learn to recognize and avoid WCR larvae, and will thereby reduce their contribution to WCR suppression. In controlled feeding assays, the effectiveness of WCR haemolymph defences varied across three predator taxa (crickets, centipedes, and ants). Centipedes (Chilopoda: Lithobiidae) were minimally affected by WCR defences, but crickets [Gryllus pennsylvanicus Burmeister (Orthoptera: Gryllidae)] spent less time feeding on WCR than on an undefended control prey, house fly maggots. However, we uncovered no evidence indicating that experienced crickets rapidly learn to avoid WCR larvae, indicating that haemolymph defences offer few, if any, survival benefits for WCR. Colonies of ants [Lasius neoniger Emery (Hymenoptera: Formicidae)] switched from low worker participation in initial attacks on WCR to higher worker participation in subsequent attacks, indicating an attempt to overcome, rather than avoid, WCR haemolymph defences. These results suggest that a diverse assemblage of natural enemies will show a diverse array of behavioural responses to toxic pest prey, and highlight the importance of behavioural diversity in driving the function of natural enemy assemblages.     

Greenhouse-Selected Resistance to Cry3Bb1-Producing Corn in Three Western Corn Rootworm Populations

Transgenic corn producing the Bacillus thuringiensis (Bt) toxin Cry3Bb1 has been useful for controlling western corn rootworm, Diabrotica virgifera virgifera LeConte, one of the most economically important crop pests in the United States. However, rapid evolution of resistance by this beetle to Bt corn producing Cry3Bb1 has been reported previously from the laboratory, greenhouse, and field. Here we selected in the greenhouse for resistance to Cry3Bb1 corn in three colonies of WCR derived from Kansas, Minnesota, and Wisconsin, respectively. Three generations of rearing on Cry3Bb1 corn significantly increased larval survival on Cry3Bb1 corn, resulting in similar survival in the greenhouse for selected colonies on Cry3Bb1 corn and isoline corn that does not produce Bt toxin. After four to seven generations of rearing on Cry3Bb1 corn, survival in the field on Cry3Bb1 corn relative to isoline corn more than doubled for selected colonies (72%) compared with control colonies (33%). For both selected and control colonies, survival in the field was significantly lower on Cry3Bb1 corn than on isoline corn. On isoline corn, most fitness components were similar for selected colonies and control colonies. However, fecundity was significantly lower for selected colonies than control colonies, indicating a fitness cost associated with resistance. The rapid evolution of resistance by western corn rootworm to Bt corn reported here and previously underlines the importance of effective resistance management for this pest.     

Characterization of western corn rootworm (Coleoptera: Chrysomelidae) population dynamics in relation to landscape attributes

Abstract 1 The western corn rootworm (WCR), Diabrotica virgifera virgifera Leconte (Coleoptera: Chrysomelidae), creates economic and environmental concerns in the Corn Belt region of the U.S.A. To supplement the population control tactics of the Areawide Pest Management Program in Brookings, South Dakota, Geographical Information Systems (GIS) were used to examine the spatial relationships from 1997 to 2001 between WCR population dynamics, habitat structure, soil texture and elevation. 2 Using the inverse distance weighted interpolation technique, WCR population density maps were created from georeferenced emergence and post‐emergence traps placed in maize fields. For each year, these maps were overlaid with vegetation, soil and elevation maps to search for quantitative relationships. 3 Through visual interpretation and correlation analysis, shifts in landscape structure, such as size, number and arrangement of patches, were shown to associate with WCR population abundance and distribution in varying degrees. Contingency analysis showed that WCR population abundance is associated with soil texture and elevation. 4 An understanding of the interactions between WCR population dynamics and landscape variables provides information to pest managers, and this can be used to identify patterns in the landscape that promote high insect population density patches to improve pest management strategies.     

Microbial Communities Associated with the Larval Gut and Eggs of the Western Corn Rootworm

Background

The western corn rootworm (WCR) is one of the economically most important pests of maize. A better understanding of microbial communities associated with guts and eggs of the WCR is required in order to develop new pest control strategies, and to assess the potential role of the WCR in the dissemination of microorganisms, e.g., mycotoxin-producing fungi.

Methodology/Principal Findings

Total community (TC) DNA was extracted from maize rhizosphere, WCR eggs, and guts of larvae feeding on maize roots grown in three different soil types. Denaturing gradient gel electrophoresis (DGGE) and sequencing of 16S rRNA gene and ITS fragments, PCR-amplified from TC DNA, were used to investigate the fungal and bacterial communities, respectively. Microorganisms in the WCR gut were not influenced by the soil type. Dominant fungal populations in the gut were affiliated to Fusarium spp., while Wolbachia was the most abundant bacterial genus. Identical ribosomal sequences from gut and egg samples confirmed a transovarial transmission of Wolbachia sp. Betaproteobacterial DGGE indicated a stable association of Herbaspirillum sp. with the WCR gut. Dominant egg-associated microorganisms were the bacterium Wolbachia sp. and the fungus Mortierella gamsii.

Conclusion/Significance

The soil type-independent composition of the microbial communities in the WCR gut and the dominance of only a few microbial populations suggested either a highly selective environment in the gut lumen or a high abundance of intracellular microorganisms in the gut epithelium. The dominance of Fusarium species in the guts indicated WCR larvae as vectors of mycotoxin-producing fungi. The stable association of Herbaspirillum sp. with WCR gut systems and the absence of corresponding sequences in WCR eggs suggested that this bacterium was postnatally acquired from the environment. The present study provided new insights into the microbial communities associated with larval guts and eggs of the WCR. However, their biological role remains to be explored.