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The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, has evolved resistance to nearly every management tactic utilized in the field. This study investigated the resistance mechanisms in a WCR strain resistant to the Bacillus thuringiensis (Bt) protein eCry3.1Ab using dsRNA to knockdown WCR midgut genes previously documented to be associated with the resistance. ATP-binding cassette transporter (ABCC4), aminopeptidase-N, cadherin, and cathepsin-B were previously found to be differentially expressed in eCry3.1Ab-resistant WCR larvae when compared to susceptible larvae after feeding on maize expressing eCry3.1Ab and its near-isoline. Here we compared the susceptibility of resistant and susceptible WCR larvae to eCry3.1Ab protein in presence or absence of dsRNA targeting the above genes using 10-day diet overlay toxicity assays. Combining ABCC4 dsRNA with eCry3.1Ab protein increased susceptibility to Bt protein in WCR-resistant larvae, but the other three genes had no such effect. Among 65 ABC transport genes identified, several were expressed differently in resistant or susceptible WCR larvae, fed on eCry3.1Ab-expressing maize versus its isoline, that may be involved in Bt resistance. Our findings provide strong evidence that ABCC4 is indirectly involved in WCR resistance to eCry3.1Ab protein by enhancing the effects of Bt-induced toxicity., Competing Interests: Competing interests: Adriano Pereira is an employee of RNAiSSANCE AG, St. Louis, MO, USA. All other authors declare no competing interests. Ethics approval and consent to participate: The experimental research and collection of plant and insect materials of this study comply with the relevant institutional, national, and international guidelines and legislation., (© 2024. The Author(s).)

The northern corn rootworm, Diabrotica barberi Smith & Lawrence (Coleoptera: Chrysomelidae) is a major pest of maize in the United States Corn Belt. Recently, resistance to Bacillus thuringiensis (Bt) maize was reported in North Dakota and increased use of Bt maize hybrids could facilitate resistance evolution in other maize-producing states. In this study, susceptibility to Bt proteins was evaluated in wild D. barberi populations from 8 fields collected in 5 different states (Minnesota, Missouri, Nebraska, Iowa, and North Dakota). Field populations were compared to a susceptible D. barberi colony in seedling and diet toxicity assays conducted with 3 concentrations of Cry3Bb1 (0.4, 4.0, and 40.0 µg/cm2) and Gpp34/Tpp35Ab1 (previously called Cry34/35Ab1; 1.4, 14.0, and 140.0 µg/cm2). The 2019 population from Meeker Co., Minnesota (MN-2019), exhibited the lowest mortality to Cry3Bb1 and also had nominally lowest mortality to Gpp34/Tpp35Ab1 at the highest concentrations tested in diet toxicity assays. Percent second instar was also highest for larvae of the Minnesota population surviving Cry3Bb1. In seedling assays, MN and IA-2018 populations exhibited the highest proportion survival and dry weight to both proteins expressed in corn. No significant differences in mortality, percent second instar, and dry weight were observed at the highest concentration for both proteins among the populations collected in in 2020. Most D. barberi populations were still highly susceptible to Cry3Bb1 and Gpp34/Tpp35Ab1 proteins based on diet and seedling assays, but resistance appears to be developing in some D. barberi populations. Now that methods are available, resistance monitoring may also be needed for D. barberi in some regions., (Published by Oxford University Press on behalf of Entomological Society of America 2023.)

The development of artificial diets could considerably simplify and reduce the cost of mass rearing of natural enemies compared to conventional rearing methods. However, improvement of artificial diets can be tedious, convoluted and often uncertain. For accelerating diet development, a better method that can offer informative feedback to target deficiencies in diet improvement is required. Our previous research demonstrated several biological characteristics were diminished in the insect predator, Arma chinensis Fallou, fed on an artificial diet formulated with the aid of transcriptomic methods compared to the Chinese oak silk moth pupae. The present study reports differential proteomic analysis by iTRAQ-PRM, which unravels the molecular mechanism of A. chinensis responding to improvements in the artificial diet. Our study provides multivariate proteomic data and provides comprehensive sequence information in studying A. chinensis . Further, the physiological roles of the differentially expressed proteins and pathways enable us to explain several biological differences between natural prey-fed and improved diet-fed A. chinensis , and subsequent proposed reformulation optimizations to artificial diets.

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), poses a serious threat to maize (Zea mays L.) growers in the U.S. Corn Belt. Transgenic corn expressing Bacillus thuringiensis (Bt) Berliner is the major management tactic along with crop rotation. Bt crops targeting WCR populations have been widely planted throughout the Corn Belt. Rootworms have developed resistance to nearly all management strategies including Bt corn. Therefore, there is a need for new products that are not cross-resistant with the current Bt proteins. In this study, we evaluated the susceptibility of WCR strains resistant and susceptible to Cry3Bb1 to the biological insecticide Spear-T (GS-omega/kappa-Hexatoxin-Hv1a) alone and combined with Cry3Bb1 protein. The activity of Hv1a alone was similar between Cry3Bb1-resistant and susceptible strains (LC50s = 0.95 mg/cm2 and 1.50 mg/cm2, respectively), suggesting that there is no cross-resistance with Cry3Bb1 protein. Effective concentration (EC50), molt inhibition concentration (MIC50), and inhibition concentration (IC50) values of Hv1a alone were also similar between both strains, based on non-overlapping confidence intervals. Increased mortality (64%) was observed on resistant larvae exposed to Hv1a (0.6 mg/cm2) + Cry3Bb1 protein (170.8 µg/cm2) compared to 0% mortality when exposed to Cry3Bb1 alone and 34% mortality to Hv1a alone (0.3 mg/cm2). The time of larval death was not significantly different between Hv1a alone (3.79 mg/cm2) and Hv1a (0.6 mg/cm2) + Cry3Bb1 (170.8 µg/cm2). New control strategies that are not cross-resistant with current insecticides and Bt proteins are needed to better manage the WCR, and Hv1a together with Cry3Bb1 may fit this role., (© The Author(s) 2021. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The western corn rootworm (WCR), Diabrotica virgifera LeConte, is the most serious pest of maize in the United States. In pursuit of developing a diet free of antibiotics for WCR, we characterized effects of thermal exposure (50-141 °C) and length of exposure on quality of WCRMO-2 diet measured by life history parameters of larvae (weight, molting, and survival) reared on WCRMO-2 diet. Our results indicated that temperatures had non-linear effects on performance of WCRMO-2 diet, and no impacts were observed on the length of time exposure. The optimum temperature of diet processing was 60 °C for a duration less than 30 min. A significant decline in development was observed in larvae reared on WCRMO-2 diet pretreated above 75 °C. Exposing WCRMO-2 diet to high temperatures (110-141 °C) even if constrained for brief duration (0.9-2.3 s) caused 2-fold reduction in larval weight and significant delays in larval molting but no difference in survival for 10 days compared with the control diet prepared at 65 °C for 10 min. These findings provide insights into the effects of thermal exposure in insect diet processing.

Mass production of Coenosia attenuata Stein at low cost is very important for their use as a biological control agent. The present study reports the performance of C. attenuata adults when reared on Drosophila melanogaster Meigen or Bradysia impatiens (Johannsem). Different densities (6, 9, 15, 24 and 36 adults per predator) of D. melanogaster or (6, 12, 24, 36 and 48 adults per predator) of B. impatiens were used at 26 ± 1 °C, 14:10 (L:D) and 70 ± 5% RH. The results concluded that C. attenuata adults had higher fecundity, longer longevity and less wing damage when reared on B. impatiens adults compared to D. melanogaster adults. Additionally, C. attenuata adults demonstrated greater difficulty catching and carrying heavier D. melanogaster adults than lighter B. impatiens adults. In this case, 12 to 24 adults of B. impatiens daily per predator were considered optimal prey density in the mass rearing of adult C. attenuata .

The northern corn rootworm (NCR), Diabrotica barberi Smith & Lawrence, is an economic pest of maize in the U.S. Corn Belt. The objective of this study was to determine the baseline susceptibility of a laboratory NCR strain to Bt proteins eCry3.1Ab, mCry3A, Cry3Bb1, and Cry34/35Ab1 using seedling, single plant, and diet-toxicity assays. Plant assays were performed in greenhouse using corn hybrids expressing one of the Bt proteins and each respective near-isoline. Diet-toxicity assays, consisting of Bt proteins overlaid onto artificial diet were also conducted. In both plant assays, significantly more larvae survived Cry34/35Ab1-expressing corn compared with all other Bt-expressing corn, and larvae that survived eCry3.1Ab-expressing corn had significantly smaller head capsule widths compared with larvae that survived Cry34/35Ab1-expressing corn. In seedling assays, larvae surviving eCry3.1Ab-expressing corn also had significantly smaller head capsule widths compared with larvae that survived mCry3A-expressing corn. Additionally, larvae that survived mCry3A-expressing corn weighed significantly more than larvae surviving eCry3.1Ab- and Cry34/35Ab1-expressing corn. In single plant assays, no significant differences in larval dry weight was observed between any of the Bt-expressing corn. In diet assays, LC50s ranged from 0.14 (eCry3.1Ab) to 10.6 µg/cm2 (Cry34/35Ab1), EC50s ranged from 0.12 (Cry34/35Ab1) to 1.57 µg/cm2 (mCry3A), IC50s ranged from 0.08 (eCry3.1Ab) to 2.41 µg/cm2 (Cry34/35Ab1), and MIC50s ranged from 2.52 (eCry3.1Ab) to 14.2 µg/cm2 (mCry3A). These results establish the toxicity of four Bt proteins to a laboratory diapausing NCR strain established prior to the introduction of Bt traits and are important for monitoring resistance evolution in NCR field populations., (© The Author(s) 2020. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Predators and pathogens often regulate the population dynamics of their prey or hosts. When species interact with both their predators and their pathogens, understanding each interaction in isolation may not capture the system's dynamics. For instance, predators can influence pathogen transmission via consumptive effects, such as feeding on infected prey, or non-consumptive effects, such as changing the prey's susceptibility to infection. A prey species' infection status can, in turn, influence predator's choice of prey and have negative fitness consequences for the predator. To test how intraguild predation (IGP), when predator and pathogen share the same prey/host, affects pathogen transmission, predator preference, and predator fitness, we conducted a series of experiments using a crop pest (Pseudoplusia includens), a generalist predator (Podisus maculiventris), and a generalist pathogen (Autographa californica multicapsid nuclear polyhedrovirus, AcMNPV). Using a field experiment, we quantified the effects of consumptive and non-consumptive predators on pathogen transmission. We found that a number of models provided similar fits to the data. These models included null models showing no effects of predation and models that included a predation effect. We also found that predators consumed infected prey more often when choosing between live infected or live healthy prey. Infected prey also reduced predator fitness. Developmental times of predators fed infected prey increased by 20% and longevity decreased by 45%, compared with those that consumed an equivalent number of non-infected prey. While this research shows an effect of the pathogen on intraguild predator fitness, we found no support that predators affected pathogen transmission.

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, is an important pest of maize (Zea mays L.). Published WCR diets contain corn root powder, which is not available for purchase, thereby limiting the practical use of diets containing this ingredient. We applied response surface modeling combined with mixture designs to formulate a WCR diet that does not require corn root powder. We developed the new formulation by systematically exploring eight protein ingredients from animal, plant, and yeast sources based on simultaneous evaluation of three life history parameters (weight, molting, and survival). This formulation (WCRMO-2) without corn root powder supported approximately 97% of larval survival and successful molting. Larval weight gain after 10 days of feeding on WCRMO-2 was 4-fold greater than that of larvae feeding on the current best published WCR diet. Additionally, there was no significant difference in these larval performance traits when larvae were reared on WCRMO-2 and the best proprietary WCR diet. A commercial version of WCRMO-2 was tested and found to perform comparably for these traits. These improvements met our goal of a diet comprised of available ingredients that supports performance of WCR larvae equal to or better than publicly available formulations and proprietary formulations.

Western corn rootworm (Diabrotica virgifera virgifera LeConte) is a serious pest of maize (Zea mays L.) in North America and parts of Europe. With most of its life cycle spent in the soil feeding on maize root tissues, this insect is likely to encounter and interact with a wide range of soil and rhizosphere microbes. Our knowledge of the role of microbes in pest management and plant health remains woefully incomplete, yet that knowledge could play an important role in effective pest management strategies. For this study, insects were reared on maize in soils from different locations. Insects from two different laboratory colonies (a diapausing and a non-diapausing colony) were sampled at each life stage to determine the possible core bacteriome. Additionally, soil was sampled at each life stage and resulting bacteria were identified to determine the possible contribution of soil to the rootworm bacteriome, if any. We analyzed the V4 hypervariable region of bacterial 16S rRNA genes with Illumina MiSeq to survey the different species of bacteria associated with the insects and the soils. The bacterial community associated with insects was significantly different from that in the soil. Some differences appear to exist between insects from non-diapausing and diapausing colonies while no significant differences in community composition existed between the insects reared on different soils. Despite differences in the bacteria present in immature stages and in male and female adults, there is a possible core bacteriome of approximately 16 operational taxonomic units (i.e., present across all life stages). This research may provide insights into Bt resistance development, improved nutrition in artificial rearing systems, and new management strategies.

The susceptibility of western corn rootworm, Diabrotica virgifera virgifera LeConte, larvae to nine insecticides from five different classes and to Bt proteins eCry3.1Ab and mCry3A in the presence or absence of feeding stimulants, was estimated in filter paper and diet toxicity assays, respectively. The use of a synthetic feeding stimulant blend of the sugars glucose, sucrose, and fructose plus linoleic acid at a ratio of 30:4:4:0.3 mg/ml of distilled water was evaluated to determine whether they increase the efficacy of insecticides and Bt proteins. The efficacy of thiamethoxam diluted in solutions with feeding stimulants was significantly increased when compared to thiamethoxam dilutions in water (>60-fold). Differences in the efficacy of the other insecticide classes when diluted in feeding stimulant solutions were no greater than fivefold when compared to the insecticides diluted in water. The presence of corn root juice as a natural feeding stimulant diminished toxicity of the insecticides, except for thiamethoxam, even though larval fresh weight was higher when fed on root juice compared to feeding stimulant or water. The use of feeding stimulants in diet toxicity assays did not enhance efficacy of eCry3.1Ab nor mCry3A proteins. Feeding stimulants can be recommended in combination with thiamethoxam to increase larval mortality. These results are discussed in terms of applicability of feeding stimulants to improve susceptibility of western corn rootworm larvae to pesticides in general., (Published by Oxford University Press on behalf of Entomological Society of America 2019.)

The northern corn rootworm (NCR), Diabrotica barberi Smith & Lawrence, is a major pest of maize (Zea mays L.). This pest has developed resistance to insecticides and adapted to crop rotation and may already be in the early stages of adaptation to toxins produced by Bacillus thuringiensis (Bt). Toxicity bioassays using artificial diet have proven to be valuable for monitoring resistance in many species, but no artificial diet has been developed specifically for NCR larvae. Toward this end, we first evaluated known Diabrotica diets to identify a starting media. We then developed a specialized diet for NCR using an iterative approach. Screening designs including 8 diet components were performed to identify the principal nutritional components contributing to multiple developmental parameters (survival, weight, and molting). We then applied mixture designs coupled with response surface modeling to optimize a blend of those components. Finally, we validated an improved NCR diet formulation that supports approximately 97% survival and molting, and a 150% increase in larval weight after 10 days of feeding compared with the best previously published artificial diet. This formulation appears suitable for use in diet bioassays as a tool for evaluating the resistance of NCR populations to insecticides.

The western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is an important economic pest of maize (Zea mays L.) in North America and Europe. Previous efforts to formulate an artificial diet for western corn rootworm larvae highlighted an important role of corn root powder, which had a significant positive impact on several larval developmental traits. Unfortunately, this ingredient is not available for purchase. Toward the goal of developing an artificial diet for western corn rootworm larvae with all ingredients readily accessible, we conducted research to isolate essential growth factors for larval development from corn root powder to improve the performance of diet without corn root powder. For all experiments, multiple life history parameters (survival, weight, and molting) were recorded from 15-d diet bioassays. Corn roots may contain factors that assist in larval growth, but some of these factors were not fully extracted by methanol and remained in the extracted root. Methanolic extracts significantly increased molting to second instar, but did not significantly increase survival, dry weight, or molting to third instar, suggesting the primary corn root substituents affecting these factors cannot be extracted or other extraction methods may be required to extract the essential factors from corn roots. We showed that whole corn root powder was best when used in combination with all the other nutrient sources in the published western corn rootworm formulation. Corn root powder made from proprietary seed and Viking seed has similar value., (© The Author(s) 2019. Published by Oxford University Press on behalf of Entomological Society of America.)

An artificial diet formulated for continuous rearing of the predator Arma chinensis was inferior to natural prey when evaluated using life history parameters. A transcriptome analysis identified differentially expressed genes in diet-fed and prey-fed A. chinensis that were suggestive of molecular mechanisms underlying the nutritive impact of the artificial diet. Changes in the diet formulation were made based on the transcriptome analysis and tested using life history parameters. The quantity of pig liver, chicken egg, tuna fish, biotin, nicotinamide, vitamin B6, thiamine, riboflavin, vitamin C, L-glutamine, and sucrose was reduced, and wheat germ oil, calcium pantothenate and folic acid were increased. Ecuadorian shrimp was added as a partial substitute for tuna fish. Several parameters improved over six generations, including increased egg viability, and decreased egg and adult cannibalism. Additionally, several parameters declined, including longer developmental times for 2nd-5th instars, and decreased nymphal weights. The improvements in life history parameters support the use of transcriptome analyses to help direct formulation improvements. However, the decline in some parameters suggests that additional information, e.g., proteomic data, may be useful as well to maximize diet formulations.

The western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is considered the most important maize (Zea mays L.) pest in the U.S. Corn Belt. Bioassays testing susceptibility to Bacillus thuringiensis Berliner (Bt) and other toxins of corn rootworm larvae often rely on artificial diet formulations. Successful bioassays on artificial diet for corn rootworm have sometimes been challenging because of microbial contamination. Toward the long-term goal of developing a universal artificial diet for western corn rootworm larvae, we compared larval survival, dry weight, and percentage of molt in 10-d bioassays from six current diets of which we were aware. In addition, as part of longer term rearing efforts, we recorded molting over an extended period of development (60 d). Six different artificial diets, including four proprietary industry diets (A, B, C, and D), the first published artificial diet for western corn rootworm (Pleau), and a new diet (WCRMO-1) were evaluated. Western corn rootworm larval survival was above 90% and contamination was 0% on all diets for 10 d. Diet D resulted in the greatest dry weight and percentage molting when compared with the other diets. Although fourth-instar western corn rootworm larvae have not been documented previously (only three instars have been previously documented), as many as 10% of the larvae from Diet B molted into a fourth instar prior to pupating. Overall, significant differences were found among artificial diets currently used to screen western corn rootworm. In order for data from differing toxins to be compared, a single, reliable and high-quality western corn rootworm artificial diet should eventually be chosen by industry, academia, and the public as a standard for bioassays.

Larval Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae) were exposed to seven different entomopathogenic nematode species to test their potential infectivity in a laboratory setting. Known D. virgifera-infecting nematode species Heterorhabditis bacteriophora Poinar, Heterorhabditis megidis Poinar, Jackson & Klein, Steinernema feltiae Filipjev, and Steinernema carpocapsae Weiser were tested in a concerted experiment alongside Steinernema diaprepesi Nguyen & Duncan, Steinernema riobrave Cabanillas, Poinar & Raulston, and a Missouri wild-type H. bacteriophora which have not been previously tested on D. virgifera. The species S. rarum Doucet was tested separately for D. virgifera infectivity. Third-instar D. virgifera were exposed to either 60 or 120 nematodes per larva for 6 d. Following exposure, mortality was recorded and larvae were examined to determine the presence of active nematode infections. Results indicated a significantly higher proportion of larvae with active infections from the Heterorhabditidae species and S. diaprepesi than the other Steinernematidae species for both exposure rates; mortality data indicated a similar trend. Steinernema rarum showed almost no infectivity in laboratory experiments.

Insect resistance to transgenic crops is a growing concern for farmers, regulatory agencies, the seed industry, and researchers. Since 2009, instances of field-evolved Bt resistance or cross resistance have been documented for each of the four Bt proteins available for western corn rootworm (WCR), a major insect pest. To characterize resistance, WCR populations causing unexpected damage to Bt maize are evaluated in plant and/or diet toxicity assays. Currently, it is not possible to make direct comparisons of data from different Bt proteins due to differing proprietary artificial diets. Our group has developed a new, publicly available diet (WCRMO-1) with improved nutrition for WCR larvae. For the current manuscript, we tested the compatibility of all Bt proteins currently marketed for WCR on the WCRMO-1 diet and specific proprietary diets corresponding to each toxin using a susceptible colony of WCR. We also tested WCR colonies selected for resistance to each protein to assess the ability of the diet toxicity assay to detect Bt resistance. The WCRMO-1 diet is compatible with each of the proteins and can differentiate resistant colonies from susceptible colonies for each protein. Our diet allows researchers to monitor resistance without the confounding nutritional differences present between diets.

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, is the most serious insect pest of corn (Zea mays L.) in the United States and parts of Europe, and arguably one of the world's most expensive pests to control. Several diet formulations are currently used by industry and public researchers to evaluate WCR larvae in diet-toxicity bioassays. However, a publicly available diet that produces normative insects that are physiologically similar to WCR larvae reared on corn roots will accelerate development of management technologies. We report a new diet formulation that supports improved weight gain, larval development and survival compared with the only public diet for WCR that is currently available in the refereed literature. The formulation was created by using response surface methods combined with n-dimensional mixture designs to identify and improve the formulation of key ingredients. Weight gain increased two-fold, and survival and molting rates increased from 93% and 90%, respectively when reared on the public diet, to approximately 99% for both survival and molting at 11 days when reared on our new formulation. This new formulation provides a standardized growth medium for WCR larvae that will facilitate comparison of research results from various working groups and compliance with regulatory requirements.

Life-table analysis yielded demographic parameter values that indicate that Tenebrio molitor (L.) pupae are potentially more suitable factitious prey to mass-produce the predator Podisus maculiventris (Say) and are more suitable prey than the larvae. P. maculiventris developed faster (23.2 vs. 25.5 d), weighed more (females 80.9 vs. 66.6 mg and males 64.7 vs. 53.7 mg), and had a higher survival rate (0.88 vs. 0.7), fecundity, and reproductive output (87.1 vs. 22.8 eggs/female) when reared on pupae compared with larvae of T. molitor. The total protein content and soluble protein content were significantly higher in pupae (60.2 and 23%, respectively) than larvae (53.1 and 14.4%, respectively). Lipid content was significantly lower in pupae (32.1%) than larvae (35.9%), and larvae had more polyunsaturated fatty acids (83.6 vs. 56.6 mg/g) and less oleic (0.1 mg/g) and steric (6.1 mg/g) acids than pupae (37.3 and 12.3 mg/g, respectively). The total sugar content was not significantly different between pupae and larvae. However, larvae had significantly more fructose than pupae, but pupae had more galactose, glucosamine, glucose, mannose, and trehalose than larvae. Differences in nutritional composition and its impact on predator demographic parameters are potential factors that make the pupal stage a better food source.

The impact of a zoophytogenous, insect-free artificial diet and a secondary prey, pupae of Chinese oak silk moth Antheraea pernyi (Guérin-Méneville) (Lepidoptera: Saturniidae), on the developmental rate, life history parameters, and fertility was examined for F6, F9, and F12 consecutive generations for domesticated Arma chinensis (Fallou) (Heteroptera: Pentatomidae). This study showed that when fed an insect-free artificial diet during both the nymphal and adult stages, developmental times were prolonged, and fecundity, egg viability, net reproductive rates (R0), and intrinsic rates of increase (rm) declined. As a result, the cost to rear A. chinensis on the artificial diet approached 1.7 times the cost of rearing A. chinensis on pupae of A. pernyi. Future diet improvements should attempt to reduce developmental time, increase fecundity, and egg viability and use less costly nutrient sources., (© The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Increased metal availability in the environment can be detrimental for the growth and development of all organisms in a food web. In part, this toxicity is due to biotransfer or bioaccumulation of metals between trophic levels. We evaluated the survival, growth, and development of a generalist Hemipteran predator (Podisus maculiventris) when fed herbivorous prey (Spodoptera exigua) reared on artificial diet amended with Cu, Zn, Ni, and Co. Predator nymphs were fed S. exigua larvae raised on diet amended with sublethal (Minimum Sublethal Concentration or MSC) or lethal (Minimum Lethal Concentration or MLC) concentrations of each metal, as well as control diet. We determined if metals were biotransferred or bioaccumulated from the diet to herbivore and predator, as well as if predator growth or survival was affected by herbivore diet. Podisus maculiventris fed herbivores raised on MLC levels of both Cu and Zn took significantly longer to mature to adults, whereas their overall survival was not affected by prey diet metal concentration for any metal. Adult weights were significantly reduced for predators raised on herbivores reared on diets amended with the MLC of Cu and Zn. Copper and Zn were bioaccumulated from diet to herbivore and from herbivore to predator, whereas Ni was biotransferred (although concentrations decreased as trophic level increased). The pattern for Co was more complex, with biotransfer the main outcome. Our results show that availability of metals in a food web can affect growth and development of a hemipteran predator, and that metals are transferred between trophic levels, with metal-specific biotransfer and bioaccumulation outcomes.

Background: The insect predator, Arma chinensis, is capable of effectively controlling many pests, such as Colorado potato beetle, cotton bollworm, and mirid bugs. Our previous study demonstrated several life history parameters were diminished for A. chinensis reared on an artificial diet compared to a natural food source like the Chinese oak silk moth pupae. The molecular mechanisms underlying the nutritive impact of the artificial diet on A. chinensis health are unclear. So we utilized transcriptome information to better understand the impact of the artificial diet on A. chinensis at the molecular level., Methodology/principal Findings: Illumina HiSeq2000 was used to sequence 4.79 and 4.70 Gb of the transcriptome from pupae-fed and artificial diet-fed A. chinensis libraries, respectively, and a de novo transcriptome assembly was performed (Trinity short read assembler). This resulted in 112,029 and 98,724 contigs, clustered into 54,083 and 54,169 unigenes for pupae-fed and diet-fed A. chinensis, respectively. Unigenes from each sample's assembly underwent sequence splicing and redundancy removal to acquire non-redundant unigenes. We obtained 55,189 unigenes of A. chinensis, including 12,046 distinct clusters and 43,143 distinct singletons. Unigene sequences were aligned by BLASTx to nr, Swiss-Prot, KEGG and COG (E-value <10(-5)), and further aligned by BLASTn to nt (E-value <10(-5)), retrieving proteins of highest sequence similarity with the given unigenes along with their protein functional annotations. Totally, 22,964, 7,898, 18,069, 15,416, 8,066 and 5,341 unigenes were annotated in nr, nt, Swiss-Prot, KEGG, COG and GO, respectively. We compared gene expression variations and found thousands of genes were differentially expressed between pupae-fed and diet-fed A. chinensis., Conclusions/significance: Our study provides abundant genomic data and offers comprehensive sequence information for studying A. chinensis. Additionally, the physiological roles of the differentially expressed genes enable us to predict effects of some dietary ingredients and subsequently propose formulation improvements to artificial diets.

Changes in essential dietary components alter global gene expression patterns in animals. We reported on a proteomics study designed to identify molecular markers of deficiencies in culture media developed for the oriental fruit fly, Bactrocera dorsalis. In that study, we found significant changes in expression of 70 proteins in adults of larvae reared on media lacking wheat germ oil (WGO), compared to media supplemented with WGO. Of these, a gene encoding an insect chitin-binding protein was expressed at about 120-fold higher levels in adult males reared on media supplemented with WGO. We inferred it may be feasible to develop the gene as a molecular marker of dietary lipid deficiency. The work was focused, however, on analysis of 11 day old adults. We have no information on expression of the chitin-binding protein, nor on any other proteins at other adult ages. In this paper we address the idea that the whole animal proteome changes dynamically with age. We reared separate groups of fruit fly larvae on media with and without WGO supplementation and analyzed protein expression in adult males and females age 0, 4, 8 and 12 days old using 2D electrophoresis. Gel densitometry revealed significant increases (by >2-fold) and decreases (by >50%) in expression levels of 29 proteins in females and 10 in males. We identified these proteins by mass spectrometry on MALDI TOF/TOF and bioinformatic analyses of the protein sequences. Two proteins, peroxiredoxin (26-fold increase) and vitellogenin 1 (15-fold increase) increased in expression in day 8 females. The key finding is that most changes in protein expression occurred in day 8 females. We infer that the fruit fly proteome changes with adult age. The natural changes in proteome with adult age is a crucial aspect of developing these and other proteins into molecular markers of lipid deficiency in fruit flies and possibly other insect species., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Ecological research requires large samples for statistical validity, typically hundreds or thousands of individuals, which are most efficiently gathered by mass-collecting techniques. For the study of interspecific interactions, molecular gut-content analysis enables detection of arthropod predation with minimal disruption of community interactions. Field experiments have demonstrated that standard mass-collection methods, such as sweep netting, vacuum sampling and foliage beating, sometimes lead to contamination of predators with nontarget DNA, thereby compromising resultant gut-content data. We deliberately contaminated immature Coleomegilla maculata and Podisus maculiventris that had been fed larvae of Leptinotarsa decemlineata by topically applying homogenate of the alternate prey Leptinotarsa juncta. We then attempted to remove contaminating DNA by washing in ethanol or bleach. A 40-min wash with end-over-end rotation in 80% EtOH did not reliably reduce external DNA contamination. Identical treatment with 2.5% commercial bleach removed most externally contaminating DNA without affecting the detectability of the target prey DNA in the gut. Use of this bleaching protocol, perhaps with minor modifications tailored to different predator-prey systems, should reliably eliminate external DNA contamination, thereby alleviating concerns about this possible source of cross-contamination for mass-collected arthropod predators destined for molecular gut-content analysis., (Published 2012. This article is a US Government work and is in the public domain in the USA.)

Fumigant activity of 34 commercial essential oils was assessed on female adults and eggs of twospotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae) at three temperatures (5, 15, and 25 degrees C). Common thyme, cinnamon, and lemongrass oils were equally effective on twospotted spider mite adults showing 85.8-100% mortality at 5 and 10 microl/liter air at 25 degrees C. At a lower temperature of 15 degrees C, lemongrass and peppermint resulted in > or =90% mortality of adults at 10 microl/liter air. Only lemongrass was relatively active at 5 microl/liter air, at 15 degrees C. At 5 degrees C, lemongrass and peppermint caused significantly higher adult mortality than controls but only at 10 microl/liter air. Common thyme oil showed the highest ovicidal activity at 5 microl/liter air at 25 degrees C. Among the main components of common thyme and lemongrass oils, citral was lethal to twospotted spider mite adults at all tested temperatures. Carvacrol, thymol, and citral caused the same inhibitory effects on the hatch of twospotted spider mite eggs at 25 degrees C. However, citral was more active than other compounds to twospotted spider mite eggs at 15 degrees C. Therefore, we conclude that citral has the best potential for development as a fumigant against twospotted spider mite on agricultural products harvested late in the growing season.

Changes in animal nutrition, particularly essential dietary components, alter global gene expression patterns. Our goal is to identify molecular markers that serve as early indicators of the quality of insect culture media. Markers of deficient culture media will increase the efficiency of developing optimal systems for mass rearing beneficial insects and some pest species because decisions on culture media quality can be made without waiting through one or several life cycles. The objective of our current study is to discover molecular markers of essential dietary lipid deficiency in the oriental fruit fly, Bactrocera dorsalis. We reared groups of fruit flies separately on media either devoid of or supplemented with wheat germ oil (WGO) and analyzed gene expression in third instar larvae and F(1) eggs using 2D electrophoresis. Gel densitometry revealed significant changes in expression levels of genes encoding eight proteins in larvae and 22 proteins in eggs. We identified these proteins by using mass spectrometry (MALDI TOF/TOF) and bioinformatic analyses of the protein sequences. Among these, we identified one gene encoding the receptor of activated C Kinase 1 (RACK1) that increased in expression by 6.8-fold in eggs from adults that were reared as larvae on media supplemented with WGO. RACK1 is an essential component of at least three intracellular signal transduction pathways, making it a good molecular marker candidate of lipid deficiency in fruit flies and possibly many other insect species., (© 2010 Wiley Periodicals, Inc.)

Culture medium supplemented with wheat germ oil (WGO) causes physiological reactions, such as increased fecundity and mobility, in some insects. Although the impact of WGO on insect physiology is important, the mechanisms of these actions are poorly understood. In this paper, we test the hypothesis that the addition of WGO to medium developed for larval oriental fruit flies modulates gene expression in the corresponding adults. We separately reared larvae of Bactrocera dorsalis on diets lacking or supplemented with WGO, and analyzed for expressed proteins in the resulting adult males and females by 2D-electrophoresis. Analysis of the gels revealed significant changes in expression levels of >70 proteins, 64 of which were identified by mass spectrometric analysis on MALDI-TOF/TOF. Apparent changes in expression levels for 6 of these proteins were confirmed by quantitative real-time PCR, showing that the changes in mRNA expression were reflected in changes in protein expression. These findings support the hypothesis that one mechanism of WGO actions in insect nutrition is the modulation of gene expression., (Copyright (c) 2010. Published by Elsevier Ltd.)

Cues that elicit a characteristic localized search behavior by neonate larvae of the western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), were extracted from living corn, Zea mays L., roots with acetone. Larvae were exposed to corn roots or to an acetone extract of corn roots and then transferred into a bioassay arena where their movements were tracked and recorded. After a 5-min exposure to live corn roots, larvae produced highly convoluted tracks that were indicative of a localized search behavior, and these distinctive tracks were also produced by larvae exposed to an acetone extract of corn roots. Larvae exposed to a filter paper control moved in relatively straight paths that were indicative of ranging behavior. Larval tracks were recorded by means of a videocamera and tracking software, and four parameters of movement were used to quantify the tracks: mean turn angle, mean meander, total distance, and maximum distance from origin. For every parameter measured, tracks resulting from exposure to the control were significantly different from tracks resulting from exposure to live corn roots and to all doses of the corn root extract. In a separate experiment, larvae exposed to corn root pieces or corn root juice exhibited the localized search behavior, but larvae exposed to oat root pieces and oat root juice (nonhost) exhibited ranging behavior.

Homalodisca vitripennis Germar 1821 (Hemiptera: Cicadellidae) [Takiya et al. (2006) Ann Entomol Soc Am 99:648-655; syn. H. coagulata (Say)] salivary gland and gut EST libraries were used to isolate cDNA fragments of the gene transcripts encoding for cathepsin L, asparaginyl endopeptidase, cathepsin B, metalloendopeptidase, cathepsin D, multicatalytic endopeptidase, and a sugar-binding C-type lectin. Transcript levels were evaluated in immature and adult H. vitripennis feeding on sunflower (Helianthus annuus) or cowpea (Vigna unguiculata). Northern blot hybridization results showed that expression of most of the transcripts were similar for all developmental stages and feeding on the two diets examined. However, the expression of the transcript for asparaginyl endopeptidase was less expressed in sunflower-fed adult females compared to sunflower-fed immatures. Also, the expression of the C-type lectin transcript was up-regulated in adults compared to immatures when fed on either diet. Documenting both the presence and variation of transcript expression involved in putative digestive proteolysis in this xylem-feeding leafhopper is noteworthy and aids efforts to design specific diet formulations for mass production of hosts and parasitoids to be used as effective biological control measures.

Three grass host species--tall fescue, Festuca arundinacea Schreber; meadow fescue, Festuca pratensis Hudson; and perennial ryegrass, Lolium perenne L.--each infected with a number of different Neotyphodium endophyte isolates, were investigated for their effects on fall armyworm, Spodoptera frugiperda (J.E. Smith). Alkaloid profiles varied among associations. Choice and no-choice tests comparing feeding and early development of S. frugiperda larvae on endophyte-infected and endophyte-free leaf blade material were performed. Endophyte-mediated resistance to S. frugiperda was greatest in meadow fescue and weakest in tall fescue. Some endophyte isolates, particularly in perennial ryegrass and meadow fescue, had a major effect on feeding and development of S. frugiperda, whereas others had no effect or were only weakly efficacious. In tall fescue, some associations deterred S. frugiperda from feeding in choice tests but had no effect on development, whereas larvae reared on other associations weighed significantly more than control larvae fed endophyte-free grass. It was concluded that the deleterious consequences of endophyte infection were easily masked by other factors in tall fescue. Relative leaf age had no effect on feeding preferences in the three host species. Chemical analysis of herbage from the plants used, and results from a no-choice study using spiked artificial diets, failed to individually implicate any of the major known alkaloids (peramine, lolitrem B, ergovaline, and lolines) in the observed effects on S. frugiperda. Hypotheses explaining these observations, and their impact on creating desirable grass-endophyte associations for use in pastures, are discussed.

The tarnished plant bug, Lygus hesperus Knight, is a pest that causes considerable economic losses to vegetables, cotton, canola, and alfalfa. Detailed knowledge of its digestive physiology will provide new opportunities for a sustainable pest management approach to control this insect. Little is known about the different protease class contributions to the overall digestion of a specific protein. To this end, the proteolytic activities in female adult L. hesperus salivary gland and midgut homogenates were quantified over a range of pH's and time points, and the contribution of different classes of proteases to the degradation of FITC-casein was determined. In the salivary gland, serine proteases were the predominant class responsible for caseinolytic activity, with the rate of activity increasing with increasing pH. In contrast, both aspartic and serine proteases contributed to caseinolytic activity in the midgut. Aspartic protease activity predominated at pH 5.0 and occurred immediately after incubation, whereas serine protease activity predominated at pH 7.5 after a 9h delay and was resistant to aprotinin. The salivary serine proteases were distinctly different from midgut serine proteases, based on the tissue-specific differential susceptibility to aprotinin and differing pH optima. Collectively, the caseinolytic activities complement one another, expanding the location and pH range over which digestion can occur.

Fragments of two artificial diet up-regulated and two prey up-regulated transcripts were isolated from the predatory Pentatomid Perillus bioculatus using suppression subtractive hybridization. A BlastX search found similarities for two diet-upregulated clones, i.e., the tyrosine-3-monooxygenase gene and the gene for the chitin binding protein, Gasp. The probe generated from the tyrosine-3-monooxygenase clone hybridized to two transcripts 2.3 and 1.2kb in size. The two transcripts were differentially regulated: the 2.3kb transcript was upregulated in the first and late third instar diet-fed nymphs, whereas the 1.2kb transcript was upregulated in the second and early third instar diet-fed nymphs. The Gasp gene was upregulated in late third instar nymphs. A positive correlation was found between levels of expression of the isolated genes and the number of generations the insects had been reared on the artificial diet.

The braconid wasp Toxoneuron nigriceps induced qualitative and quantitative changes in the protein composition of the moth Heliothis virescens host hemolymph. Total protein concentration was found to be higher in parasitized host 4 days after parasitism as compared to control hosts, mainly due to changes in a particular group of proteins. Host proteins with a molecular mass of 173 and 72 kDa were found in higher levels in the hemolymph of parasitized larvae as control hosts approached pupation, while an 80 kDa peptide was found in reduced concentration in the hemolymph of parasitized hosts. Levels of these three peptides were maintained throughout parasitoid development, while two of them (173 and 72 kDa) were cleared from the host hemolymph close to pupation. Besides the regulation of host proteins, three parasitism-specific proteins (PSPs) were released into the host hemolymph. Two of them (PSP1-MW=116 kDa, pI=6.3; PSP2-MW=114 kDa, pI=6.2) first appeared in the hemolymph of parasitized hosts soon after pupation of control host and increased in concentration as the parasitoid developed. The third PSP (PSP3-MW=56 kDa, pI=5.8) was produced towards the end of parasitoid larval development, close to parasitoid egression. Database searches based on the amino acid composition and amino terminal sequence of PSP1 and PSP2 did not produce any significant matches, while PSP3 was identified as a putative chitinase. Incubation of host derived tissues, parasitoid larvae and teratocytes in 35S conditioned media suggested PSPs were a product of teratocytes. The role of the regulation of host proteins and release of PSPs by teratocytes for the successful development of T. nigriceps are discussed.

The effect of juvenile hormone (JH) III on the hemolymph composition of vitellogenin was examined in Perillus bioculatus. Adult females were treated topically with JH III, and the premature presence of vitellogenin in the hemolymph was then detected using electrophoresis and Western blot analyses. JH III treatment resulted in a dose-dependent early production of vitellogenin that was detectable 48 h before vitellogenin was present in non-treated insects. Vitellogenin was not observed in the hemolymph of JH III-treated adult males. The techniques reported here may be useful for the detection, isolation and characterization of compounds with JH-like activity in P. bioculatus and other species of Heteroptera (which are thought to have JH-like substances other than the JHs with known chemical identity). These same techniques may also provide a method for researchers to investigate the interactions of JH-like compounds and other substances, such as ecdysteroid, in the regulation of vitellogenesis in Heteroptera.

Selected morphological and physiological properties of the corpus allatum (CA)-corpus cardiacum (CC) complex from the two-spotted stinkbug, Perillus bioculatus (Hemiptera: Pentatomidae), were studied. The CAs play an important role in insect physiology because of their production of the juvenile hormones (JHs), i.e., key hormones involved in development and reproduction. We found that the P. bioculatus CA-CC complex is present in two distinct morphological forms, the more frequently observed complex containing one "fused" CA between two CCs and the more rarely observed complex having one CA laterally attached to each CC. These complexes were tested for their ability to synthesize JH-like compounds. We found that the primary lipophilic compound synthesized by the CA-CCs migrated differently from JH III (a JH found in numerous insect species) when subjected to thin-layer chromatography. Furthermore, the synthesis of this compound is stimulated by 2E,6E-farnesol, a known precursor for JH III. These data indicate that the P. bioculatus CA- CC product has chemical properties similar to that of other (as of yet unidentified) hemipteran JHs. In addition, we found that the synthesis of this product is sensitive to pH and buffer type; minimally or not affected by the absence of the CC; expressed at similar levels in days 5-30 postemergent adults; and inhibited or decreased in adults reared under low temperature-short day conditions.

Lepidopteran larvae resist baculovirus infection by selective apoptosis of infected midgut epithelial cells and by sloughing off infected cells from the midgut. Once the infection breaches the midgut epithelial barrier and propagates from infective foci to the haemocoel, however, there are few mechanisms known to account for the resistance and clearance of infection observed in some virus-host combinations. The hypothesis that factors present in the plasma of infected pest larvae act to limit the spread of virus from initial infective foci within the haemocoel was tested. An in vitro bioassay was developed in which Helicoverpa zea single capsid nucleopolyhedrovirus (HzSNPV) was incubated with plasma collected from uninfected Heliothis virescens larvae. Infectious HzSNPV particles were then titrated on HzAM1 cells. Diluted plasma from larval Heliothis virescens exhibited a virucidal effect against HzSNPV in vitro, reducing the TCID(50) ml(-1) by more than 64-fold (from 4.3+/-3.6x10(5) to 6.7+/-0.6x10(3)). The antiviral activity was heat-labile but was unaffected by freezing. In addition, protease inhibitors and specific chemical inhibitors of phenol oxidase or prophenol oxidase activation added to diluted plasma eliminated the virucidal activity. Thus, in the plasma of larval lepidopterans, the enzyme phenol oxidase may act as a constitutive, humoral innate antiviral immune response.

The impact of a zoophytogenous, insect-free artificial diet on the developmental rate, life history parameters, and fertility was examined over 11 consecutive generations for domesticated Perillus biocullatus (F.) (Heteroptera: Pentatomidae). This study showed that when fed an insect-free artificial diet during both the nymphal and adult stages, developmental times were prolonged, and the net reproductive rates (R0) and the intrinsic rates of increase (r(m)) were significantly lower than when fed larval Trichoplusia ni at both nymphal and adult stages. Moreover, the cost to rear P. bioculatus on the artificial diet approached 1.1 times the cost of rearing P. bioculatus on natural prey. These results demonstrate the effectiveness and potential cost-savings of the zoophytogenous artificial diet for rearing a beneficial pentatomid.

The active-toxin form of CrylAc (65 kDa) or Cry2Ab was fed to a non-susceptible insect, Lygus hesperus, in an artificial diet. Biochemical and immunocytochemical methods were used to determine the distribution of ingested toxin. The toxins did not elicit a feeding deterrent response. CrylAc and Cry2Ab were ingested; small amounts were absorbed into the hemolymph as holoproteins, but most was excreted. SDS-PAGE analysis of CrylAc and Cry2Ab incubations with salivary gland homogenate showed a small decrease in the molecular weight of the active toxins. Proteolytic processing of the toxins also occurred in vivo, within the digestive system of L. hesperus. Excreted CrylAc and Cry2Ab retained activity toward lepidopteran larvae. Immunocytochemical in vivo localization studies showed negligible association of CrylAc with L. hesperus tissues. In contrast, strong extracellular association of Cry2Ab was observed with L. hesperus midgut brush border microvilli and basement membrane, as well as with cellular outlines within the hemolymph and fat body.

The impact of a zoophytophagous, insect-free artificial diet upon the developmental rate, life table parameters, and fertility table parameters was examined over 11 consecutive generations for domesticated and wild colonies of Podisus maculiventris (Say) (Heteroptera: Pentatomidae). This study showed that the developmental time, preoviposition period, fecundity, and nymphal survival improved in the domestic colony when fed an insect-free artificial diet for 11 consecutive generations, but remained relatively constant for the wild colony, as did reproductive rate and intrinsic rate of increase. Although, after 11 generations of adaptation to an artificial diet feeding regime, all reproductive and fertility table parameters were still significantly lower than when fed on T. ni larvae as the natural prey, the realized cost of rearing either colony on the artificial diet approached 1.2 times the cost of rearing these insects on a natural prey. This is a significant achievement in the effort to develop cost-effective artificial diets for the mass-rearing of beneficial pentatomids, and has positive implications for the use of one artificial diet to efficiently rear several beneficial insects.

Selected compounds were used to study physiological processes associated with digestion in the western tarnished plant bug, Lygus hesperus Knight. Durations of passage and rates of absorption, digestion, and excretion were determined for a digestible protein (casein), a non-digestible protein (green fluorescent protein, GFP), and a non-digestible carbohydrate (dextran). Dextran was used as a control to monitor the non-absorptive flow rate of ingesta through the digestive system. Fluorescent tracking of FITC-conjugates of casein and dextran, as well as immunoblotting and immunofluorescent staining of casein and GFP, were used to monitor the degradation (in vitro) and ingestion, digestion, and distribution (in vivo) of the respective compounds. Under our experimental conditions, L. hesperus took discrete meals, feeding and excreting at 2-3 h intervals. Rate of food passage was variable. FITC-dextran was found in the fecal material of most insects by 6-8 h after treatment initiation; by 12 h, 95% of ingested FITC-dextran was recovered from all insects. FITC-casein was digested extensively in in vitro homogenates of gut, hemolymph, and salivary gland. In vivo, FITC-casein was ingested and partially absorbed as a holoprotein into the hemolymph. Ingested FITC-casein was partially degraded in the gut and hemolymph within 2 h of ingestion, and no holoprotein was found after 12 h. In contrast, there was no detectable degradation of GFP in hemolymph, gut, and salivary gland homogenates after 24 h of incubation. Ingested GFP was not degraded in gut or hemolymph up to 8 h after treatment initiation, but did transfer to the hemolymph as a holoprotein. Analysis of immunohistological images confirmed that GFP bound to gut epithelial cell brush-border membranes. However, the mechanism by which GFP and casein pass as holoproteins into the hemolymph remains unknown.

The impact of an insect-free artificial diet provided at nymphal and/or adult stage upon the developmental rate, life table parameters, and fertility table parameters was examined for Podisus maculiventris (Say). This study showed that when fed an insect-free artificial diet during both the nymphal and adult stage, developmental time was prolonged, preoviposition period was extended, and reproductive rate (R0) and intrinsic rate of increase (r) were significantly lower than when fed larval insect prey at both nymphal and adult stages. Additionally, feeding larval prey to adults reared as nymphs on an artificial diet significantly increased the proportion of fertile females, the number of eggs laid by mated females, the reproductive rate and intrinsic rate of increase, but the mean generation time was not significantly different. Likewise, feeding artificial diet to adults reared on larval prey resulted in a significant reduction in reproductive rate and intrinsic rate of increase. The "realized" cost to rear P. maculiventris on the artificial diet was calculated (as the cost to double the population size) using raw material cost, fertility table parameters and doubling time values. Raw material cost for rearing P. maculiventris colony on Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae) was only 1.4 times higher than the cost of artificial diet raw materials required to rear the same size colony. However, the realized cost of rearing was 3.5 times higher when rearing on artificial diet because of the prolonged developmental time and reduced reproductive output. The cost efficiency of rearing a beneficial insect on an artificial diet that decreases the intrinsic rate of increase of a colony is discussed, as well as the advantages and disadvantages of supplementing adult diets with natural prey at the reproductive stage.

The compatibility of the venom from the parasitic species Euplectrus comstockii Howard (Hymenoptera: Eulophidae) with the pathogenicity of Autographa californica (Speyer) (Lepidoptera: Noctuidae) MNPV baculovirus (AcMNPV) was tested in third and fourth instar larvae of Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae). The presence of AcMNPV did not alter the ability of the venom to arrest ecdysis in T. ni larvae. The presence of the venom delayed the rate of viruses by AcMNPV but increased the total mortality rates from days 9 to 14 in both third and fourth instar T. ni larvae. The delay in viruses was minimized by administering the virus prior to envenomation. In the presence of the venom, the final LD50 values were lower for fourth instar larvae than for third instar larvae. Surface response equations were developed to visualize the effect of the venom on the viruses caused by AcMNPV.

Parasitic Hymenoptera regulate their hosts in order to provide a suitable source of nutrition and dwelling for their offspring. Few regulatory factors known to cause a specific effect on the host have been structurally characterized. The larval ectoparasitoid Euplectrus comstockii Howard (Hymenoptera: Eulophidae) arrests larval-larval ecdysis in its lepidopteran hosts. Prior to oviposition, the female wasp inject a venom into the hemocoel of the host and that venom alone is effective in causing the arrestment. A venom gland-reservoir structure connected to the lower reproductive tract of the wasp contains a complex mixture of proteins. There are no obvious similarities among the electrophoretic banding pattern (native or denatured) for venom proteins of E comstockii and several other parasitic hymenopteran species. Venomous protein, separated by electrophoretic techniques, with a native mol. wt of c. 66,000, was capable of arresting larval-larval ecdysis in 4th instar larvae of Trichoplusia ni (Lepidoptera: Noctuidae). Nanogram quantities of the protein were sufficient to cause arrestment. The activity of the protein was sensitive to temperature, pH, organic solvent, and protease.

The cuticular hydrocarbons of diapausing pupae of the tobacco hornworm, Manduca sexta (L.), are composed of two types of hydrocarbons: n-alkanes (3%) and unsaturated hydrocarbons (97%). The integument, fat body, muscle, and hemolymph are composed of three types of hydrocarbons: n-alkanes (9, 6, 9, and 2%, respectively), branched alkanes (75, 85, 80, and 56%, respectively), and unsaturated hydrocarbons (16, 9, 11, and 42%, respectively). The absence of branched alkanes on the cuticular surface indicated that hydrocarbons are selectively synthesized or deposited according to the presence or absence of methyl branches in the molecule. The hydrocarbons consisted of homologous series of n-alkanes from 21 to 41 carbon atoms, monomethylalkanes from 23 to 39 carbon atoms, dimethylalkanes from 25 to 43 carbon atoms, and trimethylalkanes from 33 to 37 carbon atoms. All branched components had methyl groups near the center of the molecule, and the di- and trimethyl branched components had three methylene units between the branch points. Straight chain alkenes, alkadienes, and alkatrienes from 23 to 44 carbon atoms were partially characterized. The percentage composition of each homologous series varied from tissue to tissue.

Insect cells of an established line, Drosophila Kc cells, take up and metabolize juvenile hormone (JH). The cytoplasm of these cells contains a protein that binds JH with specificity, saturability, and high affinity (K(d) = 1.56 x 10(-8) M). The kinetics for the specific binding and dissociation of JH I were independently measured, and the rate constants were found to be k(a) = 1.3 x 10(6) M(-1) min(-1), k(d) = 1.3 x 10(-2) min(-1), respectively. All three juvenile hormones bind to the protein with comparable affinities; the corresponding acid or diol metabolites of JH I are not bound. About 2500 hormone-binding protein molecules are present per cell. The protein has a molecular weight of 80,000 as estimated by gel permeation chromatography and by sucrose gradient sedimentation. The properties of this protein suggest that it functions as a cytoplasmic receptor for juvenile hormone.