Comparative susceptibility of western corn rootworm (Coleoptera: Chrysomelidae) adults to selected insecticides in Kansas

Susceptibility of adult populations of the western corn rootworm, Diabrotica virgifera virgifera LeConte, to several insecticides was evaluated in seven Kansas counties, including Dickinson, Ford, Finney, Pottawatomie, Republic, Riley, and Stevens, between 1996 and 2002. All populations surveyed were highly susceptible to methyl parathion with the largest difference in susceptibility of only three-fold based on 16 complete bioassays for the populations from six counties over a 5-yr period. Noticeable decreases in carbaryl susceptibility were found in populations collected from Republic County between 1997 and 2001 when the cucurbitacin-carbaryl-based bait SLAM was widely used as an areawide management approach for adult corn rootworm control. However, the lowered carbaryl susceptibility returned to previous levels 1 yr after the use of SLAM was halted in the managed (treated) cornfields. This change implies possible dispersal of insects into the relatively small managed area from surrounding untreated cornfields and / or some fitness cost associated with carbaryl resistance within the population. Relative susceptibility of western corn rootworm adults also was evaluated for seven commonly used insecticides, including bifenthrin, carbaryl, chlorpyrifos, cypermethrin, fipronil, malathion, and methyl parathion. They were tested with corn rootworm adults collected from a single cornfield. Methyl parathion and bifenthrin were highly toxic to corn rootworm adults, and cypermethrin, chlorpyrifos, carbaryl, and malathion were only slightly less toxic. Although fipronil was highly toxic to adult rootworms, its activity was much slower than that of other insecticides. Thus, bifenthrin and methyl parathion were among the most effective in killing corn rootworm adults.     

Inheritance of methyl-parathion resistance in Nebraska western corn rootworm populations (Coleoptera: Chrysomelidae)

Field populations of western corn rootworm, Diabrotica virgifera virgifera LeConte, were collected from three different sites (York Co., Phelps Co., and Saunders Co.) in Nebraska during 1996. Adult bioassays of these three populations were conducted with different concentrations of methyl-parathion and at a diagnostic concentration (1.0 microg/ml) to determine resistance levels among these populations. Self and reciprocal crosses were made between the two resistant and one susceptible laboratory-reared populations. Dose-responses and dominance ratios calculated for the four reciprocal crosses indicated that resistance was incompletely dominant in both strains, although in one of the strains there was an indication of sex linkage. However, evaluation of native polyacrylamide gels stained for nonspecific esterases and nonspecific esterase activity of parents and F1 progeny of the crosses suggested that esterase inheritance was completely dominant and autosomal. The results of this study were inconclusive with regard to the precise nature of inheritance, because the bioassays and esterase assays could not discriminate between heterozygotes and homozygotes. However, they do provide insight into the potential for developing simple diagnostic assays to assess resistance frequencies. Based on the inheritance studies described in this investigation, we can begin to generate information on specific genetic factors that dictate the evolutionary divergence of discrete resistant populations and facilitate modeling efforts designed to approximate the movement of genes for resistance among populations.     

Field screening maize germplasm for resistance and tolerance to western corn rootworms (Col.: Chrysomelidae)†

Abstract: In the hopes of lessening the current reliance on soil insecticides, developing a viable alternative for transgenic maize hybrids, and providing sustainable options for Europe, researchers recently have been developing novel maize lines that exhibit resistance and/or tolerance to corn rootworm larvae. Here we report the results of a 2‐year field experiment in a northern growing region assessing the resistance and tolerance of 10 experimental synthetic maize populations selected for varying levels of damage from western corn rootworm larvae, Diabrotica virgifera virgifera LeConte (Col.: Chrysomelidae) and four maize hybrids. Maize non‐preference, antibiosis and tolerance to rootworms was evaluated using previously established methods, including: the Iowa 1–6 root damage rating scale, root fresh weight, compensatory root growth ratings and adult rootworm emergence. Among the experimental synthetic maize populations, BS29‐11‐01 was the most susceptible, and had a mean root damage rating that was greater than the highly susceptible maize hybrid B37 × H84. This line also had the lowest mean root fresh weight and one of the lowest mean compensatory root growth ratings. In contrast, CRW8‐3 appeared to be tolerant to western corn rootworms, and had the lowest mean root damage rating, which was comparable with that of the non‐transgenic hybrid DeKalb^® 46‐26.     

Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae) larval feeding behavior on transgenic maize (MON 863) and its isoline

Diabrotica species (Coleoptera: Chrysomelidae) larval behavior studies have posed a challenge to researchers because of the subterranean life cycle of this pest. To fully understand how the western corn rootworm, Diabrotica virgifera virgifera LeConte, injures the maize, Zea mays L., root system, its behavior must be studied. For example, larvae that can detect an area of the root that has a lower amount of toxin, whether from an insecticide or a transgenic maize plant, have an increased chance of survival. This study assessed D. v. virgifera larval feeding behavior on rootworm-susceptible maize and maize containing a biotechnology-derived trait (MON 863) with resistance to D. v. virgifera first instar feeding. Maize plants were grown in a medium that allowed for direct observation and measurements during feeding of larval stadia. Neonates were placed on maize seedlings, and data were taken at 3, 6, 9, and 12 d postinfestation on resistant and susceptible maize. On rootworm-susceptible maize, neonate larvae aggregated at the root tips and began actively feeding, and then they moved to older root tissue. Conversely, some larvae that ingested Cry 3Bb1 from the resistant maize exhibited no movement. Other larvae on the resistant maize moved continuously, sampling root hairs or root tissue but not actively feeding. The continuously moving larvae had visibly empty guts, suggesting possible nonpreference for the resistant root. This study contributes to our understanding of D. v. virgifera larval behavior and provides insight into questions surrounding the potential evolution of behavioral and biochemical resistance to Cry3Bb1.     

Larval sampling and instar determination in field populations of northern and western corn rootworm (Coleoptera: Chrysomelidae)

Abundance and head capsule width were measured for northern (Diabrotica barberi Smith & Lawrence) and western corn rootworm (D. virgifera virgifera LeConte) larvae recovered primarily from maize root systems but also from large soil cores each centered around a root system. Larvae for measurement derived from field populations under infestation and rotation regimes that allowed most specimens to be assigned to species. A frequency distribution of head capsule widths indicated three separate peaks for western corn rootworm, presumably representing frequency of the three larval instars, with no larvae measuring 280 or 420 microm in the valleys between peaks. Multiple normal curves fit to similar but partially overlapping peaks generated by northern corn rootworm suggested that division of first to second and second to third instar can best be made for this species at 267 and 406 microm, respectively (270 and 410 when measurements are made to the nearest 20 microm). These results implied that instar of individuals from mixed northern and western corn rootworm populations can be accurately judged from head capsule width without having to determine species. The relative abundance of western corn rootworm instars was similar in root systems removed from the center of 19-cm diameter x 19-cm deep soil cores and in soil cores from which the root systems were removed. Furthermore, the number of larvae from root systems correlated significantly with that from the surrounding soil. These results indicated that the former and much more convenient sampling unit can be used to estimate population developmental stage and possibly density, at least early in the season when these tests were done and young larvae predominated.     

Post-establishment movement of western corn rootworm larvae (Coleoptera: Chrysomelidae) in Central Missouri corn

If registered, transgenic corn, Zea mays L., with corn rootworm resistance will offer a viable alternative to insecticides for managing Diabrotica spp. corn rootworms. Resistance management to maintain susceptibility is in the interest of growers, the Environmental Protection Agency, and industry, but little is known about many aspects of corn rootworm biology required for an effective resistance management program. The extent of larval movement by the western corn rootworm, Diabrotica virgifera virgifera LeConte, that occurs from plant-to-plant or row-to-row after initial establishment was evaluated in 1998 and 1999 in a Central Missouri cornfield. Post-establishment movement by western corn rootworm larvae was clearly documented in two of four treatment combinations in 1999 where larvae moved up to three plants down the row and across a 0.46-m row. Larvae did not significantly cross a 0.91-m row after initial host establishment in 1998 or 1999, whether or not the soil had been compacted by a tractor and planter. In the current experiment, western corn rootworm larvae moved from highly damaged, infested plants to nearby plants with little to no previous root damage. Our data do not provide significant insight into how larvae might disperse after initial establishment when all plants in an area are heavily damaged or when only moderate damage occurs on an infested plant. A similar situation might also occur if a seed mixture of transgenic and isoline plants were used and if transgenic plants with rootworm resistance are not repellent to corn rootworm larvae.     

Insecticidal proteins from Bacillus thuringiensis protect corn from corn rootworms.

Field tests of corn co-expressing two new delta-endotoxins from Bacillus thuringiensis (Bt) have demonstrated protection from root damage by western corn rootworm (Diabrotica virgifera virgifera LeConte). The level of protection exceeds that provided by chemical insecticides. In the bacterium, these proteins form crystals during the sporulation phase of the growth cycle, are encoded by a single operon, and have molecular masses of 14 kDa and 44 kDa. Corn rootworm larvae fed on corn roots expressing the proteins showed histopathological symptoms in the midgut epithelium.     

Adaptation by western corn rootworm (Coleoptera: Chrysomelidae) to Bt maize: inheritance, fitness costs, and feeding preference

We examined inheritance of resistance, feeding behavior, and fitness costs for a laboratory-selected strain of western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), with resistance to maize (Zea maize L.) producing the Bacillus thuringiensis Berliner (Bt) toxin Cry3Bb1. The resistant strain developed faster and had increased survival on Bt maize relative to a susceptible strain. Results from reciprocal crosses of the resistant and susceptible strains indicated that inheritance of resistance was nonrecessive. No fitness costs were associated with resistance alleles in the presence of two entomopathogenic nematode species, Steinernema carpocapsae Weiser and Heterorhabditis bacteriophora Poinar. Larval feeding studies indicated that the susceptible and resistant strains did not differ in preference for Bt and non-Bt root tissue in choice assays.     

Estimation of efficacy functions for products used to manage corn rootworm larval injury

Maize, Zea mays L., is an economically important crop grown throughout the world. Corn rootworm, Diabrotica spp. (Coleoptera: Chrysomelidae), larvae constitute a significant economic threat to maize production in the United States, where yield losses and management costs associated with corn rootworm species exceed $1 billion annually. Furthermore, the introduction of the western corn rootworm, D. virgifera virgifera LeConte, into maize‐producing regions of Europe has made managing corn rootworm larval injury an international concern. Larvae injure maize plants by feeding on root tissue and are the primary target of management activities. Products commonly used to protect root systems from injury include chemical insecticides (seed or soil applied) and genetically modified maize hybrids expressing toxins derived from Bacillus thuringiensis Berliner (Bt). The confirmation of field‐evolved resistance to various Bt toxins in populations of the western corn rootworm presents a significant management challenge. We performed a meta‐analysis to provide a broad understanding of the relative efficacy of the primary products currently being used to manage corn rootworm larval injury, including insecticidal seed treatments, soil insecticides and Bt hybrids (with and without the addition of soil insecticide). Our analysis is unique in the breadth of locations and years included – we analysed 135 individual trials conducted from 2003 through 2014 at multiple sites in both Illinois and Nebraska. Panel data were produced by pairing the mean node‐injury rating for each treatment of a given trial with the mean node‐injury rating for untreated maize. Linear regression models were developed to estimate the relationship between the potential for corn rootworm larval injury and product performance. For a given level of injury potential, the parameters estimated reveal differences in the degree of root protection offered by the various product categories analysed. Implications for developing long‐term, integrated, and sustainable practices for managing this important pest of maize are discussed.     

Immunological assessment of an insecticide resistance-associated esterase in the Western corn rootworm

In previous investigations, we have determined that organophosphate resistance in the western corn rootworm, Diabrotica virgifera virgifera, is at least partially attributed to a group of non-specific carboxylesterases referred to as group II. Antiserum raised against a purified 66-kDa group II esterase is specific for the denatured enzyme. This antiserum reacts similarly with both beetle homogenates from resistant and susceptible populations, although there is much higher signal intensity in immunoblots of resistant relative to susceptible beetles. These results suggest that overproduction of group II esterases is the underlying basis of esterase-mediated resistance in D. v. virgifera by demonstrating that (1) group II esterases are immunologically indistinguishable between the resistant and susceptible populations, and (2) the intensity differences are due to increased group II esterase proteins in the resistant population. The diagnostic potential of immunological-based assays was tested with a traditional diagnostic concentration bioassay and a biochemical-based native PAGE assay. Significant correlations were observed among all three diagnostic assays (regression coefficients ranging from 0.95 to 0.96). These results demonstrate the importance of the 66-kDa protein as a resistance-associated biochemical marker, thus emphasizing the potential for 66-kDa protein-targeted immunoassays in resistance monitoring programs.     

Antixenosis in maize reduces feeding by western corn rootworm larvae (Coleoptera: Chrysomelidae)

SUM2162 is the first known example of a naturally occurring maize, Zea mays L., genotype with antixenosis (nonpreference) resistance to western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), larval feeding. Behavioral responses of neonate western corn rootworm larvae were evaluated in laboratory bioassays with seven maize genotypes selected for native resistance to rootworm feeding damage. Two susceptible maize genotypes and one transgenic (Bacillus thuringiensis) maize genotype were included as controls. In soil bioassays with cut roots, no larvae entered the roots of the resistant variety SUM2162, but at least 75% of the larvae entered the roots of every other maize type. Larvae made significantly fewer feeding holes in the roots of SUM2162 than in all the other maize genotypes, except the isoline control. In feeding bioassays, larval feeding varied significantly among maize genotypes, but there was no significant difference between the resistant varieties and the susceptible controls. There were no significant differences among any of the genotypes in host recognition (search) behavior of larvae after exposure to the roots. Little variation in feeding stimulant blends was observed among maize genotypes, indicating minimal contribution to the observed antixenosis.     

Western corn rootworm and Bt maize: challenges of pest resistance in the field

Crops genetically engineered to produce insecticidal toxins from the bacterium Bacillus thuringiensis (Bt) manage many key insect pests while reducing the use of conventional insecticides. One of the primary pests targeted by Bt maize in the United States is the western corn rootworm, Diabrotica virgifera virgifera LeConte. Beginning in 2009, populations of western corn rootworm were identified in Iowa, USA that imposed severe root injury to Cry3Bb1 maize. Subsequent laboratory bioassays revealed that these populations were resistant to Cry3Bb1 maize, with survival on Cry3Bb1 maize that was three times higher than populations not associated with such injury. Here we report the results of research that began in 2010 when western corn rootworm were sampled from 14 fields in Iowa, half of which had root injury to Cry3Bb1 maize of greater than 1 node. Of these samples, sufficient eggs were collected to conduct bioassays on seven populations. Laboratory bioassays revealed that these 2010 populations had survival on Cry3Bb1 maize that was 11 times higher and significantly greater than that of control populations, which were brought into the laboratory prior to the commercialization of Bt maize for control of corn rootworm. Additionally, the developmental delays observed for control populations on Cry3Bb1 maize were greatly diminished for 2010 populations. All 2010 populations evaluated in bioassays came from fields with a history of continuous maize production and between 3 and 7 y of Cry3Bb1 maize cultivation. Resistance to Cry34/35Ab1 maize was not detected and there was no correlation between survival on Cry3Bb1 maize and Cry34/35Ab1 maize, suggesting a lack of cross resistance between these Bt toxins. Effectively dealing with the challenge of field-evolved resistance to Bt maize by western corn rootworm will require better adherence to the principles of integrated pest management.     

Effect of MIR604 transgenic maize at different stages of development on western corn rootworm (Coleoptera: Chrysomelidae) in a central Missouri field environment

The establishment and survival of western corn rootworm, Diabrotica virgifera virgifera LeConte, was evaluated on transgenic Bacillus thuringiensis Berliner maize, Zea mays L., expressing the mCry3A protein (MIR604) and non-Bt maize with the same genetic background (isoline maize) at different stages of development in 2007 and 2008. Overall, western corn rootworm larval recovery, root damage, and adult emergence were significantly higher on isoline maize compared with MIR604. The number of larvae and adults collected from MIR604 did not significantly differ among egg hatch dates from each maize developmental stage evaluated in either year. In 2007, damage to isoline maize roots was lower than expected and never exceeded 0.24 nodes of damage. In 2008, over 0.60 nodes of damage occurred on isoline maize roots. The mean weight and head capsule width of larvae and adults recovered from MIR604 and isoline maize were generally not significantly different. Results are discussed in relation to insect resistance management of western corn rootworm.     

European populations of Diabrotica virgifera virgifera are resistant to aldrin,but not to methyl‐parathion

The western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is a major pest of cultivated corn in North America and has recently begun to invade Europe. In addition to crop rotation, chemical control is an important option for D. v. virgifera management. However, resistance to chemical insecticides has evolved repeatedly in the USA. In Europe, chemical control strategies have yet to be harmonized and no surveys of insecticide resistance have been carried out. We investigated the resistance to methyl‐parathion and aldrin of samples from nine D. v. virgifera field populations originating from two European outbreaks thought to have originated from two independent introductions from North America. Diagnostic concentration bioassays revealed that all nine D. v. virgifera field populations were resistant to aldrin but susceptible to methyl‐parathion. Aldrin resistance was probably introduced independently, at least twice, from North America into Europe, as there is no evident selection pressure to account for an increase of frequency of aldrin resistance in each of the invasive outbreaks in Europe. Our results suggest that organophosphates, such as methyl‐parathion, may still provide effective control of both larval and adult D. v. virgifera in the European invasive outbreaks studied.     

Belowground resistance to western corn rootworm in lepidopteran-resistant maize genotypes

Several maize, Zea mays L., inbred lines developed from an Antiguan maize population have been shown to exhibit resistance to numerous aboveground lepidopteran pests. This study shows that these genotypes are able to significantly reduce the survival of two root feeding pests, western corn rootworm, Diabrotica virgifera virgifera LeConte, and southern corn rootworm, Diabrotica undecimpunctata howardi Barber. The results also demonstrated that feeding by the aboveground herbivore fall armyworm, Spodoptera frugiperda (J. E. Smith), before infestation by western corn rootworm reduced survivorship of western corn rootworm in the root tissues of some, but not all, genotypes. Likewise, the presence of western corn rootworm in the soil seemed to increase resistance to fall armyworm in the whorl in several genotypes. However, genotypes derived from the Antiguan germplasm with genetic resistance to lepidopterans were still more resistant to the fall armyworm and both rootworm species than the susceptible genotypes even after defense induction. These results suggest that there may be intraplant communication that alters plant responses to aboveground and belowground herbivores.     

Mortality of western corn rootworm larvae on MIR604 transgenic maize roots: field survivorship has no significant impact on survivorship of F1 progeny on MIR604

Mortality of western corn rootworm, Diabrotica virgifera virgifera LeConte, larvae due to MIR604 transgenic corn, Zea mays L., expressing the modified Cry3A (mCry3A) protein relative to survivorship on corn with the same genetic background without the gene (isoline corn) was evaluated at three Missouri sites in both 2005 and 2006. We made these comparisons by using wild-type western corn rootworm at three different egg densities (6,000, 3,000, and 1,500 eggs per m) so that the role of density-dependent mortality would be known. The mortality due to the mCry3A protein was 94.88% when averaged across all environments and both years. Fifty percent emergence of beetles was delayed approximately 5.5 d. Beetles were kept alive and their progeny evaluated on MIR604 and isoline corn in the greenhouse to determine whether survivorship on MIR604 in the field for one generation increased survivorship on MIR604 in the greenhouse in the subsequent generation. There was no significant difference in survivorship on MIR604 in greenhouse assays between larvae whose parents survived isoline and larvae whose parents survived MIR604 in the field the previous generation, indicating that many susceptible beetles survived MIR604 in the field the previous season along with any potentially resistant beetles. The data are discussed in terms of rootworm insect resistance management.     

Interaction between western corn rootworm (Coleoptera: Chrysomelidae) larvae and root-infecting Fusarium verticillioides

A greenhouse experiment was conducted to evaluate the effect of soil-dwelling larvae of the western corn rootworm, Diabrotica virgifera virgifera LeConte, on infection of maize roots by the mycotoxin-producing plant-pathogenic fungus, Fusarium verticillioides (Saccardo) Nirenberg (synonym=Fusarium moniliforme Sheldon). The time and order of application of F. verticillioides and western corn rootworm were varied in three different treatments to investigate the influence of timing on root colonization of F. verticillioides and western corn rootworm larval development. Root feeding by western corn rootworm larvae increased root colonization by F. verticillioides (as determined by real-time polymerase chain reaction) up to 50-fold when a high inoculum (10(7) spores/plant) of F. verticillioides was applied before western corn rootworm eggs were added. This effect was stronger the earlier F. verticillioides was applied relative to the time of western corn rootworm egg application but was only significant for the high F. verticillioides inoculum density treatment; F. verticillioides colonization was not increased when a low F. verticillioides inoculum density (10(6) spores/plant) was applied. F. verticillioides slightly suppressed larval development in that the ratio of second- to third-instar larvae was higher in treatments with F. verticillioides than without F. verticillioides. F. verticillioides reduced western corn rootworm head capsule width when applied before or simultaneously with western corn rootworm. The results of this study are discussed focusing on conditions that favor root colonization by F. verticillioides and its influence on western corn rootworm larval development.     

Diagnostic assays based on esterase-mediated resistance mechanisms in western corn rootworms (Coleoptera: Chrysomelidae)

Resistance to methyl-parathion among Nebraska western corn rootworm, Diabrotica virgifera virgifera LeConte, populations is associated with increased hydrolytic metabolism of an organophosphate insecticide substrate. An electrophoretic method to identify resistant individuals based on the staining intensity of esterase isozymes on nondenaturing polyacrylamide gels was developed. Three groups of esterases (I, II, and III) were visible on the gels, but only group II esterase isozymes were intensified in resistant populations. A total of 26 and 31 field populations of western corn rootworms from Nebraska (in 1998 and 1999, respectively) were assessed with nondenaturing polyacrylamide gel electrophoresis (PAGE) assays and diagnostic concentration bioassays. Significant correlations were observed between the two diagnostic assays. Group II esterase isozymes provide a reliable biochemical marker for detection of methyl-parathion resistance in individual western corn rootworms and a tool for monitoring the frequency of resistant individuals in field populations.     

Field-evolved resistance to Bt maize by western corn rootworm: predictions from the laboratory and effects in the field

Crops engineered to produce insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt) provide an effective management tool for many key insect pests. However, pest species have repeatedly demonstrated their ability to adapt to management practices. Results from laboratory selection experiments illustrate the capacity of pest species to evolve Bt resistance. Furthermore, resistance has been documented to Bt sprays in the field and greenhouse, and more recently, by some pests to Bt crops in the field. In 2009, fields were discovered in Iowa (USA) with populations of western corn rootworm, Diabrotica virgifera virgifera LeConte, that had evolved resistance to maize that produces the Bt toxin Cry3Bb1. Fields with resistant insects in 2009 had been planted to Cry3Bb1 maize for at least three consecutive years and as many as 6years. Computer simulation models predicted that the western corn rootworm might evolve resistance to Bt maize in as few as 3years. Laboratory and field data for interactions between western corn rootworm and Bt maize indicate that currently commercialized products are not high-dose events, which increases the risk of resistance evolution because non-recessive resistance traits may enhance survival on Bt maize. Furthermore, genetic analysis of laboratory strains of western corn rootworm has found non-recessive inheritance of resistance. Field studies conducted in two fields identified as harboring Cry3Bb1-resistant western corn rootworm found that survival of western corn rootworm did not differ between Cry3Bb1 maize and non-Bt maize and that root injury to Cry3Bb1 maize was higher than injury to other types of Bt maize or to maize roots protected with a soil insecticide. These first cases of field-evolved resistance to Bt maize by western corn rootworm provide an early warning and point to the need to apply better integrated pest management practices when using Bt maize to manage western corn rootworm.     

Characterization of general esterases from methyl parathion-resistant and -susceptible populations of western corn rootworm (Coleoptera: Chrysomelidae)

A consistent correlation between elevated esterase activity and methyl parathion resistance among Nebraska western corn rootworm, Diabrotica virgifera virgifera LeConte, populations has previously been documented. Characterization of general esterase activity using naphtholic esters as model substrates indicated that differences between resistant and susceptible strains could be maximized by optimizing assay conditions. The optimal conditions identified here were similar to those reported for other insect species. The majority of general esterase activity was found in the cytosolic fractions of resistant populations, whereas the activity was more evenly distributed between cytosolic and mitochondrial/nuclear fractions in the susceptible population. General esterase activity was predominately located in the adult thorax and abdomen. Although there were significant differences in general esterase activities between resistant and susceptible populations, the differences exhibited in single beetle activity assays did not provide sufficient discrimination to identify resistant individuals. In contrast, single larva activity assays provided greater discrimination and could be considered as an alternative to traditional bioassay techniques.