Coleopteran Insects Research Articles

English

Laccases, member of multicopper oxidase (MCO) family enzymes, play crucial roles in insects’ cuticle tanning and pigmentation. The purpose of this study was to identify and characterize a laccase gene from the red palm weevil (RPW), Rhynchophorus ferrugineus. The isolated RPW laccase gene sequence was 3,389 bp, including a 2,163 bp open reading frame that encodes 720 amino acids. The RPW laccase gene conserved the MCO functional motifs Type-3, Type-1, and Type-2, respectively. Phylogenetic analysis categorized this protein into the functional cluster 2 of insect laccases (Lac2). The primary transcripts for R. ferrugineus laccase 2 (RfeLac2) were highly expressed in the adult’s cuticle, elytra, and hindwings, and in the larval cuticles four days before molting. Then, the transcripts were declined drastically in the larval cuticles three days before molting. This implies that the suppression of RfeLac2 in the larvae occurs earlier than expected. RfeLac2 transcripts were very low in the gut and adipose tissues of larvae and adults, irrespective to the span to undergo molting. This suggests that RfeLac2 is not active in the tissues that do not undergo heavy sclerotization. Three-dimensional (3-D) structure modeling of RfeLac2 predicted eight histidines, one glycine, and one phenylalanine, as copper-binding ligands on the laccase active center. The study finding indicates that the pattern of the RPW RfeLac2 expression varies from other coleopteran insects, a phenomenon that requires further investigation.   Key words: Cuticle, expression, laccase, phylogeny, Red palm weevil.

In-vitro evaluation of antimicrobial and insect repellent potential of supercritical-carbon dioxide (SCF-CO2) extracts of selected botanicals against stored product pests and foodborne pathogens.

In the present study, the antimicrobial and the insect repellent activity of 16 botanical extracts obtained by supercritical CO2 (SCF) extraction were evaluated. The present investigation was conducted as there is a necessity for exploration of natural botanical extracts that target both stored product insects and microbes. The antimicrobial activity was studied by disc diffusion and broth microdilution methods against ten microbial species, including Gram-positive bacteria (Staphylococcus aureus, Bacillus subtilis and Listeria monocytogenes), Gram-negative bacteria (Escherichia coli and Salmonella enterica), and fungi (Aspergillus flavus, Aspergillus paraciticus, Aspergillus ochraceous, Aspergillus niger and Penicillium verrucosum). Repellency assay was carried out by area preference method against three coleopteran insects (Tribolium castaneum, Rhyzopertha dominica and Sitophilus oryzae). Among all the extracts, thyme and ajwain were effective against all the tested bacteria with a minimum inhibition concentration (MIC) of 256-1024µg/mL. Hop extract resulted in better antibacterial activity against all the tested Gram-positive bacteria with a MIC of 32-64µg/mL. Oregano, thyme and ajwain extracts showed broad-spectrum antifungal activity against all the tested fungi with MIC of 128-1024µg/mL. Most of the extracts exhibited class V (80.1-100%) repellency against T. castaneum. Extracts of hop, ajwain and thyme were found to have strong repellency against T. castaneum and R. dominica. Therefore, SCF extracts of ajwain and thyme can be explored further for the application of bio-extracts as a growth limiting factors in a microcosm where such consortia thrive.

Carob pests in the Mediterranean region: bio-ecology, natural enemies and management options

An overview on the bio-ecology, natural enemies and pest contol tools of the main pests infesting carob tree (Ceratonia siliqua L.), pods and seeds in the Mediterranean region is provided herein. Most of the species are occasionally present on host carobs where they did not cause serious damage. In particular, the occurrence of mealybugs, soft and armored scales and also mites is usually sporadic. This is also the case for dipteran and lepidopteran pests, except for the most damaging and economic species, the carob moth Ectomyelois ceratoniae (Zeller), a highly polyphagous insect that can infest carob either in open-field or in storage. Cause of wounding are bark and wood-boring coleopteran insects, which can induce severe wilting and/or death of carob branches and trunk. The recent introduction in Europe of two species of Xylosandrus spp. (X. compactus and X. crassiusculus) is of increasing importance, due to their proven initial aggressiveness in Italy. Brief details regarding moth and rodent populations residing inside the storage buildings are also emphasized. Obviously, one aspect warranting special attention concerns the introduction and rapid spread in the Mediterranean region of invasive alien insect species such as some ambrosia beetles.

Identification and characterization of multiple dsRNases from a lepidopteran insect, the tobacco cutworm, Spodoptera litura (Lepidoptera: Noctuidae)

RNA interference (RNAi) efficiency varies among insects. RNAi is highly efficient and systemic in coleopteran insects but quite variable and inefficient in lepidopteran insects. Degradation of double-stranded RNA (dsRNA) by double-stranded ribonucleases (dsRNases) is thought to contribute to the variability in RNAi efficiency observed among insects. One or two dsRNases involved in dsRNA digestion have been identified in a few insects. To understand the contribution of dsRNases to reduced RNAi efficiency in lepidopteran insects, we searched the transcriptome of Spodoptera litura and identified six genes coding for DNA/RNA non-specific endonucleases. Phylogenetic analysis revealed the evolutionary expansion of dsRNase genes in insects. The mRNA levels of three midgut-specific dsRNases increased during the larval stage, and the highest dsRNA-degrading activity was detected in third-instar larvae. Proteins produced via the expression of three midgut-specific dsRNases, and the widely expressed dsRNase3, in a baculovirus system showed dsRNase activity for four out of five dsRNases tested. In addition, the increase in dsRNA-degrading activity and upregulation of dsRNase1 and 2 in larvae fed on cabbage leaves suggests that the diet of S. litura can influence dsRNase expression, dsRNA stability, and thus probably RNAi efficiency. This is the first report that multiple dsRNases function together in an RNAi-recalcitrant insect. The data included in this paper suggest that multiple dsRNases coded by the S. litura genome might contribute to the lower and variable RNAi efficiency reported in this and other lepidopteran insects.

Proteomic insights into the immune response of the Colorado potato beetle larvae challenged with Bacillus thuringiensis

Bacillus thuringiensis (Bt) toxins constitute effective, environmentally safe biopesticides. Nevertheless, insects' tolerance to Bt is influenced by environmental factors affecting immunity. To understand larval immune response in the devastating coleopteran insect pest Colorado potato beetle (CPB), we undertook a proteomic analysis of hemolymph of non-treated control larvae and larvae consuming non-lethal doses of spore-crystal mixtures containing the coleopteran-active Cry3Aa toxin. Results revealed lower amount of proteins involved in insect growth and higher amount of immune response-related proteins in challenged insects, sustaining the larval weight loss observed. Additionally, we found a potential regulatory role of the evolutionary conserved miR-8 in the insect's immune response relying on antimicrobial peptides (AMPs) production. Upon toxin challenge, different patterns of hemolymph AMPs expression and phenoloxidase activity were observed in CPB larvae reared on different Solanaceae plants. This suggests that diet and diet-associated insect midgut microbiota might modulate this insects' tolerance to non-lethal doses of Bt.

Development of Anncaliia algerae (Microsporidia) in Drosophila melanogaster

Representatives of the genus Anncaliia are known as natural parasites of dipteran and coleopteran insects, amphipod crustaceans, but also humans, primarily with immunodeficiency. Anncaliia algerae-caused fatal myositis is considered as an emergent infectious disease in humans. A. (=Nosema, Brachiola) algerae, the best studied species of the genus, demonstrates the broadest among microsporidia range of natural and experimental hosts, but it has never been propagated in Drosophila. We present ultrastructural analysis of development of A.algerae in visceral muscles and adipocytes of Drosophila melanogaster 2wk after per oral experimental infection. We observed typical to Anncaliia spp. features of ultrastructure and cell pathology including spore morphology, characteristic extensions of the plasma membrane, and presence of "ridges" and appendages of tubular material at proliferative stages. Anncaliia algerae development in D.melanogaster was particularly similar to one of A.algerae and A.(Brachiola) vesicularum in humans with acute myositis. Given D.melanogaster is currently the most established genetic model, with a fully sequenced genome and easily available transgenic forms and genomic markers, a novel host-parasite system might provide new genetic tools to investigate host-pathogen interactions of A.algerae, as well to test antimicrosporidia drugs.

Spermatogenic Alterations in the Ground Beetle Trachyderma hispida (Coleoptera: Tenebrionidae) Induced by Ceramic Industrial Pollution

A coleopteran insect, Trachyderma hispida, was collected from the proximity of ceramic factories in Khorshed district, Alexandria, Egypt, to elucidate the testicular damage elicited by heavy metal pollution. Bioaccumulation of heavy metals was estimated by using X-ray microanalysis. Histological and ultrastructure examination revealed various changes in germ cells in insects collected from the polluted region. The inducible histological alterations included: occlusion and collapsed follicles, disrupted cyst wall and necrotic spermatogenic elements. Ultrastructure changes implied abnormal chromatin condensation, ill-defined nuclear membrane, vacuolated cytoplasm, disintegrated nebenkern, deterioration of mitochondrial derivatives and degeneration of axonemes. In addition, agglutinated spermatids, and sperms with a double tail were noticed. The most noteworthy feature in spermatogonia and spermatocyte was the intra-tubular inclusion bodies. Histological and electron micrographs pointed out the structure of the parietal cell which functioned as a Sertoli cell. Adverse damage to these cells by ceramic pollution was reported. In conclusion, by using insects as bioindicators, ceramic production may impair reproduction.

Effects of Bacillus thuringiensis Genetic Engineering on Induced Volatile Organic Compounds Emission in Maize and the Attractiveness to a Parasitic Wasp.

In order to control lepidopteran and coleopteran insects, the genes expressing Bacillus thuringiensis (Bt) insecticidal proteins have been transferred into crops. Ecological risk assessments of the transgenic plants have included impacts on non-target entomophagous insects, such as parasitoid wasps. Herbivore-induced plant volatiles are considered to be important defensive traits of plants because these compounds play as an important role in recruitment of natural enemies. Here, we evaluated induced volatile emissions of maize seedlings of two Bt cultivars (5422Bt1, event Bt11 and 5422CBCL, event Mon810), and their nearly isogenic non-Bt line 5422. We damaged plants mechanically and then applied with the regurgitant of Spodoptera litura (F.) caterpillars (Lepidoptera: Noctuidae), or treated the plants with the plant hormone jasmonic acid (JA), to trigger similar defensive responses of plants. Compared to the non-Bt isoline 5422 and the Bt maize 5422CBCL, the other Bt maize 5422Bt1 released more (3E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) when they were all treated by artificial wounds and caterpillar regurgitant; and released more linalool, DMNT and (E)-β-farnesene when applied with JA solution. As a result, the total volatile emission of the 5422Bt1 was highest. However, the difference in volatile emission did not affect the attractiveness of the Bt maize plants to the egg parasitoid Trichogramma ostriniae Pang et Chen (Hymenoptera: Trichogrammatidae) compared to the nearly isogenic non-Bt plants. The variability of induced volatiles of maize cultivars derived from conventional breeding programs and transgenic methods are discussed.

Metarhizium humberi sp. nov. (Hypocreales: Clavicipitaceae), a new member of the PARB clade in the Metarhizium anisopliae complex from Latin America

A new species, Metarhizium humberi, from the M. anisopliae complex and sister lineage of the M. anisopliae s.str. in the PARB clade, including M. pingshaense, M. anisopliae, M. robertsii and M. brunneum, is described based on phylogenetic analyses [translation elongation factor 1-alpha (5'TEF and 3'TEF), RNA polymerase II largest subunit (RPB1a), RNA polymerase II second largest subunit (RPB2a) and β-tubulin (BTUB)]. Metarhizium humberi was first collected in 2001 in the Central Brazilian state of Goiás, later found to be a common fungus in soils in Brazil, and since then has also been isolated from coleopteran, hemipteran and lepidopteran insects in Brazil and Mexico. This new species, named in honor of Richard A. Humber, a well-known insect pathologist and taxonomist of entomopathogenic fungi, is characterized by a high insecticidal activity against different developmental stages of arthropod pests with importance in agriculture and vectors of diseases to human and animals.

Seasonality of Insect Succession and Dog Carcass Decomposition in Different Habitats

The different collected insect species were distributed on the carcasses according to the decomposition stages or postmortem intervals (PMI). In the winter season, the insect succession showed that carcasses placed outdoor attracted the greatest numbers and highest diversity of insect species. The calliphorid fly; Chrysomya albiceps was first attracted to the bloated and decay stages. In both habitats (outdoor and indoor), the numbers and diversity of insect species on carcasses were decreased during the advanced decay stage and then increased again during the dry stage. In the spring season, Chrysomya albiceps was the first fly attracted to the fresh or bloated stage of dog carcass in both habitats. Moreover, calliphorid flies were disappeared during the advanced decay stage in both habitats. While Coleopteran insects appeared during the decay and advanced decay stages. In the summer season, the forensic insect species showed nearly the same distribution of the different decomposition stages. Also, Chrysomya albiceps was the first fly attracted to the early stages of decomposition. In general, it appeared that the diversity and numbers of forensic insect species, which colonize dog carcass, were increased outdoor and decreased indoor. In the autumn season, carcasses placed outdoor attracted the greater numbers and higher diversity of forensic insects. The calliphorid fly, Chrysomya albiceps,and housefly, Musca domestica were first attracted to the fresh and bloated stages, especially of carcasses placed outdoor. The adult beetles, Dermestes maculatus, Necrobia rufipes and Hister sp.were early seen during bloated, decay and advanced decay stages.

Isoenzymes and protein polymorphism in Blaps polycresta and Trachyderma hispida (Forsskål, 1775) (Coleoptera: Tenebrionidae) as biomarkers for ceramic industrial pollution.

The expression levels of Esterase Isoenzyme and total soluble protein fractionation were studied in two coleopteran insects Blaps polycresta and Trachyderma hispida to evaluate the possible hazards from ceramic and plastic factories in the Khorshed Region, East of Alexandria, Egypt. Two insect collection sites were selected. The first site was the garden of the Faculty of Science, Moharram Bek, Alexandria University, which is considered a non-polluted site, and Khorshed district, considered as the polluted site. Percentages of heavy metals were estimated using SEM-X-ray microanalysis of soft tissues of both sexes of the two coleopteran insects. Esterase Isoenzymes were found to be overexpressed in B. Polycresta but not T. hispida. Female B. polycresta from the polluted site exhibited overexpression of the second and third loci. Furthermore, the females were found to be more affected than males, which only showed the overexpression of the second loci. T. hispida (females and males) collected from the reference site were found to have increased esterase activity compared with those sampled from the polluted site. The Snake-Skin™ Dialysis tubing technique, used for optimizing the protein extraction method, reflected the highest quantified proteins compared to other, traditional methods. SDS polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the whole-body protein reflected definite variations between T. hispida and B. polycresta in fraction number and activity at the two sites. Varied expression levels for metallothionein (MT) heavy metal resistance proteins for B. polycresta and T. hispida were also detected in the study. Based on these results, we suggest that biochemical biomarkers could infer environmental hazards, B. polycresta and T. hispida are successful biomarkers for heavy metal pollution.

TmToll-7 Plays a Crucial Role in Innate Immune Responses Against Gram-Negative Bacteria by Regulating 5 AMP Genes in Tenebrio molitor.

Although it is known that the Drosophila Toll-7 receptor plays a critical role in antiviral autophagy, its function in other insects has not yet been reported. Here, we have identified a Toll-like receptor 7 gene, TmToll-7, in the coleopteran insect T. molitor and examined its potential role in antibacterial and antifungal immunity. We showed that TmToll-7 expression was significantly induced in larvae 6 h after infection with Escherichia coli and Staphylococcus aureus and 9 h after infection with Candida albicans. However, even though TmToll-7 was induced by all three pathogens, we found that TmToll-7 knockdown significantly reduced larval survival to E. coli, but not to S. aureus, and C. albicans infections. To understand the reasons for this difference, we examined the effects of TmToll-7 knockdown on antimicrobial peptide (AMP) gene expression and found a significant reduction of E. coli-induced expression of AMP genes such as TmTenecin-1, TmDefensin-1, TmDefensin-2, TmColeoptericin-1, and TmAttacin-2. Furthermore, TmToll-7 knockdown larvae infected with E. coli showed significantly higher bacterial growth in the hemolymph compared to control larvae treated with Vermilion dsRNA. Taken together, our results suggest that TmToll-7 plays an important role in regulating the immune response of T. molitor to E. coli.

My Times with Professor Igor Kregar and some Hydrolytic Enzymes

The late Professor Igor Kregar and this author had several overlapping biochemical research interests. One focus was the glycoside hydrolase, lysozyme, and the other was proteolytic enzymes and their inhibitors, in particular those present in tissues from insects and plants. Regarding lysozyme our results helped to understand its catalytic mechanism and the carboxylic acid functional groups involved. Another area was insect-plant interactions involving defensive responses of plants to insect feeding via proteolytic enzyme inhibitors. Those results can be utilized in transgenic plant and seed biotechnological applications, which would help to reduce damage to plants and seeds caused by coleopteran and other insect pests. Also described are some cultural and travel interactions that the author benefited from his friendship with Professor Kregar.

My Times with Professor Igor Kregar and some Hydrolytic Enzymes.

The late Professor Igor Kregar and this author had several overlapping biochemical research interests. One focus was the glycoside hydrolase, lysozyme, and the other was proteolytic enzymes and their inhibitors, in particular those present in tissues from insects and plants. Regarding lysozyme our results helped to understand its catalytic mechanism and the carboxylic acid functional groups involved. Another area was insect-plant interactions involving defensive responses of plants to insect feeding via proteolytic enzyme inhibitors. Those results can be utilized in transgenic plant and seed biotechnological applications, which would help to reduce damage to plants and seeds caused by coleopteran and other insect pests. Also described are some cultural and travel interactions that the author benefited from his friendship with Professor Kregar.

Vibrations in hemipteran and coleopteran insects: behaviors and application in pest management

Many groups of insects utilize substrate-borne vibrations for communication. They display various behaviors in response to vibrations in sexual and social communication and in predator–prey interactions. Although the number of reports on communication and behaviors using vibrations has continued to increase across various insect orders, there are several studies of the exploitation of vibrations for pest management in Hemiptera and Coleoptera. Here, we review the studies of behaviors and communication using vibrations in hemipteran and coleopteran insects. For instance, pentatomid bugs display species- and sex-specific vibrational signals during courtship, whereas cerambycid beetles show startle responses to vibrations in the context of predator–prey interactions. Concepts and case studies in pest management using vibrations—especially regarding the disruption of communication and behavior—are also presented.

Identification of cytochrome P450 monooxygenase genes and their expression in response to high temperature in the alligatorweed flea beetle Agasicles hygrophila (Coleoptera: Chrysomelidae)

Cytochrome P450 monooxygenases (P450s) are a large class of enzymes that play essential roles in metabolic processes such as hormone synthesis and the catabolism of toxins and other chemicals in insects. In the present study, we identified 82 P450 genes using comprehensive RNA sequencing in the flea beetle Agasicles hygrophila, and all of the sequences were validated by cloning and sequencing. Phylogenetic analysis showed that the P450 genes in A. hygrophila fell into the mitochondrial clan, CYP2 clan, CYP3 clan and CYP4 clan and were classified into 20 families and 48 subfamilies. Most A. hygrophila P450 genes had high sequence homology with those from other coleopteran insects. To understand the effects of high temperatures on the metabolic processes of female and male adults, we studied the effects of two temperature regimes (constant temperature of 28 °C for 20 h with a 4-h period of high temperatures of 30 °C and 39 °C) on the expression levels of P450 genes in A. hygrophila using RT-PCR and qRT-PCR. The results showed that there were no differences in expression in 30 P450 genes between the control and high-temperature-treated A. hygrophila adults, while 22 P450 genes showed up-regulated expression and 19 P450 genes were down-regulated in A. hygrophila female adults after high-temperature treatment. For A. hygrophila male adults exposed to high temperatures, we found that 8 P450 genes had higher expression levels and 12 P450 genes had lower expression levels under the same conditions. The P450 genes are candidates that showed significantly different expression levels after high-temperature treatments in A. hygrophila adults, and further studies are needed to determine their possible roles in metabolic processes during the response to elevated temperatures.

Identification of a phosphine resistance mechanism in Rhyzopertha dominica based on transcriptome analysis

Abstract Worldwide, the increasing number of insecticide-resistant insects is one of the main problems in agriculture not only in the field but also in storage. The lesser grain borer, Rhyzopertha dominica, is one of the cosmopolitan insects that infests a wide range of stored cereals and related commodities. However, strongly resistant populations of R. dominica have been detected in Australia that could threaten the market access of infested commodities. A resistant strain originally collected in Australia showed extremely high levels of resistance to phosphine, around 100-fold higher than that of the susceptible strain. The resistant strain has a mutation in dihydrolipoamide dehydrogenase (DLD), which is a well-known phosphine resistance mechanism in some coleopteran insects. Thus, here, we tried to identify their phosphine resistance mechanism based on transcriptome analysis using RNA-Seq in the adult stage in addition to the mutation effect in R. dominica. Over 1 Gb was sequenced in each replication (susceptible and resistant), and similar to the synergistic test, cytochrome P450, carboxylesterase, CbE and glutathione S-transferase, GST gene families were significantly differently expressed in two strains (p

Identification of an entomopathogenic bacterium, Xenorhabdus ehlersii KSY, from Steinernema longicaudum GNUS101 and its immunosuppressive activity against insect host by inhibiting eicosanoid biosynthesis.

Steinernema longicaudum GNUS101, an entomopathogenic nematode, was isolated from soils in Korea. Its internal transcribed space sequence was highly similar to the known S. longicaudum species. Infective juveniles (IJs) of S. longicaudum were highly virulent to lepidopteran and coleopteran insects. Two different bacteria were isolated from the hemolymph of lepidopteran larvae infected with S. longicaudum. They exhibited blue and red colonies on nutrient bromothymol blue agar. The red-colored bacterium was identified as Enterococcus mundtii KHY while the blue-colored bacterium was identified as Xenorhabdus ehlersii KSY based on 16S rRNA sequencing and biochemical characters. The bacterial species showed different growth rates, with X. ehlersii KSY growing more slowly than E. mundtii KHY. Both bacteria were entomopathogenic, but showed differences in suppressing host immune responses. X. ehlersii KSY, but not E. mundtii KHY, showed inhibitory activity against cellular immune responses of Spodoptera exigua larvae including hemocyte-spreading behavior and nodule formation in bacteria-cultured broth. Its immunosuppressive activity was reversed by adding arachidonic acid, an eicosanoid biosynthesis precursor. Furthermore, organic extracts of X. ehlersii KSY using hexane or ethyl acetate showed inhibitory activity against cellular immune responses of S. exigua larvae. Arachidonic acid addition to S. exigua larvae infected with X. ehlersii significantly rescued the survival rate of target insect. Of the two bacteria isolated from S. longicaudum GNUS101, only X. ehlersii induced immunosuppression of target insect by inhibiting eicosanoid biosynthesis.

Solid cultures of thrips-pathogenic fungi Isaria javanica strains for enhanced conidial productivity and thermotolerance

Abstract Entomopathogenic fungi have great potential to control agricultural and horticultural insect pests, however optimizing conidial production systems to demonstrate high productivity and stability still needs additional efforts for successful field application and industrialization. Although many virulent entomopathogenic fungal isolates have been viewed as potential candidates in a laboratory environment, very few of the isolates are being used in practice for application in agricultural fields as commercial products. I. javanicus is an entomopathogenic fungus that is parasitic to various diverse coleopteran and lepidopteran insects and thought good candidate as biopesticdes. In this work, the basic characteristics of two entomopathogenic fungi, I. javanica FG340 and Pf04, were investigated in morphological examinations, genetic identification, and virulence against Thrips palmi, and then the feasibility of various grains substrates for conidial production was assessed, particularly focusing on conidial productivity and thermotolerance. Isaria javanica FG340 and Pf04 conidia were solid-cultured on 12 grains for 14 days in a Petri dish. Of the tested Italian millet, perilla seed, millet and barley-based cultures showed high conidial production. The four-grain media yielded >1 × 109 conidia/g of I. javanica FG340 and Pf04. Pf04 strain had enhanced thermotolerance up to 45 °C when cultured on Italian millet. In application, it was easy to make a conidial suspension using the cultured grains, and several surfactants were tested to release the conidia. This work suggests several possible inexpensive grain substrates by which to promote conidial production combined with enhanced stability against exposure to high temperature.

Double-stranded RNA binding protein, Staufen, is required for the initiation of RNAi in coleopteran insects

RNA interference (RNAi) is being used to develop methods to control pests and disease vectors. RNAi is robust and systemic in coleopteran insects but is quite variable in other insects. The determinants of efficient RNAi in coleopterans, as well as its potential mechanisms of resistance, are not known. RNAi screen identified a double-stranded RNA binding protein (StaufenC) as a major player in RNAi. StaufenC homologs have been identified in only coleopteran insects. Experiments in two coleopteran insects, Leptinotarsa decemlineata and Tribolium castaneum, showed the requirement of StaufenC for RNAi, especially for processing of double-stranded RNA (dsRNA) to small interfering RNA. RNAi-resistant cells were selected by exposing L. decemlineata, Lepd-SL1 cells to the inhibitor of apoptosis 1 dsRNA for multiple generations. The resistant cells showed lower levels of StaufenC expression compared with its expression in susceptible cells. These studies showed that coleopteran-specific StaufenC is required for RNAi and is a potential target for RNAi resistance. The data included in this article will help improve RNAi in noncoleopteran insects and manage RNAi resistance in coleopteran insects.