Comparative Study of Cowpea Storage Technologies in the Sahel Region of Niger.

Cowpea stored on smallholders' farms suffers serious losses to insect pests. A study conducted in Niger compared five postharvest technologies marketed in sub-Saharan Africa to protect stored grain. Naturally-infested cowpea stored for eight months showed adult Callosobruchus maculatus (F.) mortality of 97% to 100% in the hermetic bags (PICSTM, SuperGrainbagTM, AgroZ®, EVAL™, and ZeroFly® bags). There was no change in grain damage and weight loss of cowpea stored in hermetic bags. There was, however, a loss of up to 10 to 16% in germination when the grain was stored in hermetic bags. Results observed for grain stored in ZeroFly® bags impregnated with deltamethrin were substantial and similar to those in control woven bags. In both ZeroFly® and woven bags, (1) adult C. maculatus population augmented by 35.7% and 78.6%, (2) increased weight losses of 27.3% and 25.2%, and (3) reduced germination of 37.0% and 28.8%, respectively. After opening the bags, abrasions were noted on the liners of hermetic bags, potential damage that could limit their reuse if they only have a single liner. Smallholder farmers in the Sahel can safely store their cowpea in all the hermetic bags tested. However, further research is needed to mitigate insect damage on liners of hermetic bags to improve their performance and reusability.

Performance of Five Postharvest Storage Methods for Maize Preservation in Northern Benin.

Several postharvest technologies are currently being commercialized to help smallholder farmers in sub-Saharan Africa reduce grain storage losses. We carried out a study in Northern Benin to compare the effectiveness of five technologies being sold to protect stored grain. Maize that had been naturally infested by insects was stored in four hermetic storage technologies (SuperGrainbag™, AgroZ® bag, EVAL™, and Purdue Improved Crop Storage-PICS™ bags), an insecticide impregnated bag (ZeroFly®), and a regular polypropylene (PP) woven bag as control. Oxygen levels in hermetic bags fluctuated between 0.5 ± 0.0 (v/v) and 1.0 ± 0.3 (v/v) percent during the seven months of storage. No weight loss or insect damage was observed in grain stored in any of the hermetic storage bags after seven months. However, grain stored in ZeroFly® and PP woven bags had weight losses of 6.3% and 10.3%, respectively. These results will help farmers and development agencies when making decisions to use and/or promote storage technologies to reduce postharvest grain losses.

Effects of PICS bags on insect pests of sorghum during long-term storage in Burkina Faso.

The PICS bags, originally developed for cowpea storage, were evaluated for sorghum (Sorghum bicolor) preservation. Batches of 25 kg of sorghum grain were stored in 50 kg PICS or polypropylene (PP) bags under ambient conditions for 12 months and assessed for the presence of insect pests and their damage, seed viability and, oxygen and carbon dioxide variations. The grain was incubated for 35 days to assess whether any insects would emerge. After six months of storage, oxygen levels decreased in the PICS bags compared to polypropylene bags. After 12 months of storage, only two pests, Rhyzopertha dominica and Sitophilus zeamais were found in the PICS bags. However, in PP bags there were additional pests including Tribolium castaneum and Oryzeaphilus mercator and Xylocoris flavipes. Grain weight loss and damage caused by these insects in the PP bags were significantly higher compared to those stored in PICS bags. Germination rates of sorghum grains stored in PP bags decreased significantly while no changes were observed in grains stored in PICS bags when compared to the initial germination. After the incubation post storage period, there was a resurgence of R. dominica in sorghum grains from PICS bags but the population levels were significantly lower compared to polypropylene bags. PICS bags preserved the quality and viability of stored sorghum grains and protected it from key insect pests. The PICS technology is effective for long-term sorghum storage but the potential resurgence of insects in low-oxygen environment calls for further research.

Groundnut Production and Storage in the Sahel: Challenges and Opportunities in the Maradi and Zinder Regions of Niger.

Groundnut Arachis hypogaea (L.), is an important legume crop after cowpea Vigna unguiculata (L. Walp) in Niger. However, there has been a decline in its economic importance due to several challenges. A survey of 800 farmers was conducted in 40 villages in the Maradi and Zinder regions to assess constraints and opportunities to improve groundnut production and marketing. Average land size and yield varied by region: 1.3 ha per farmer and 461.3 kg ha-1 in Maradi, and 1.7 ha per farmer and 417.2 kg ha-1 in Zinder. Insect pests (aphids) were the most important production constraint. Groundnut is typically stored for six to eight months after harvest but 91% of farmers do not take any precautions to protect the grain. Storage enables farmers to earn high profit margins of up to 33 and 113% for unshelled and shelled groundnuts, respectively. Most farmers (71.5%) sell their groundnut in unshelled form in local and urban markets. Traders are the main buyers according to 61.7% of farmers while processors were mentioned as purchasers by less than 20%. Sales are mostly done by individual farmers while very little is sold through cooperatives. Given that groundnut is a profitable crop adapted to the Sahelian zone, there is need to improve its production, storage, and value addition through processing.

Cumulative oxygen consumption during development of two postharvest insect pests: Callosobruchus maculatus Fabricius and Plodia interpunctella Hübner.

Insect pests such as Callosobruchus maculatus Fabricius and Plodia interpunctella Hübner cause substantial losses to grain during postharvest storage. In the last few years, hermetic storage technologies have been successfully used by smallholder farmers in Africa and Asia to protect their harvested grain against insect pests. Hermetic technologies owe much of their effectiveness to restricting oxygen availability to insects confined in the containers. There is a need to better understand the biology of specific storage insect pests and their responses to hypoxia. We employed a novel and non-invasive analytical technology, the OxySense 5250i, to measure oxygen levels in closed containers, and evaluated its effectiveness in measuring the total oxygen consumption of two insect pests during their development: C. maculatus and P. interpunctella. The total amount of oxygen consumed by C. maculatus during its larval development period determined with the OxySense apparatus was not different from that previously recorded using another instrument, the Mocon Pac Check 325 gas analyzer. Using the OxySense 5250i, we found that P. interpunctella consumes nearly three times as much oxygen per insect over its larval-to-adult developmental period compared to C. maculatus. Information on the lifetime oxygen consumption of insects provides relevant information to the effectiveness and ability of hermetic technologies to protect stored products against insect pests.

A time-saving method for sealing Purdue Improved Crop Storage (PICS) bags.

Purdue Improved Crop Storage (PICS) bags were designed to reduce grain storage losses on smallholder farms. The bag consists of three layers: two high-density polyethylene liners fitted inside a woven polypropylene bag. Recently, farmer groups, development relief programs, and government food security agencies have shown interest in PICS bags for large-scale use. PICS bags are conventionally closed by a twist-tie (TT) method, which involves twisting, folding, and tying the lip of each layer individually with a cord. This is not only time and labor intensive, but also may affect the integrity of the liners. We evaluated three new bag closure methods: i) inner liner rolled onto itself and middle liner fold-tied (IR), ii) both liners folded together and tied (FT), and iii) both liners folded and tied separately (FS), along with the conventional twist tie (TT) method. The time to close partially or fully filled 50 kg-capacity PICS bags filled with maize grain was assessed. Results showed that FT was the most time-saving method, reducing bag sealing time by >34% versus the usual TT method. The average internal oxygen levels reached <2% within a week in bags containing grain highly infested with Sitophilus zeamais, while it remained >5% levels for less-infested bags. In both cases, insect population growth was suppressed. Oxygen depletion rates among tying methods remained the same regardless of the closure method used. When large numbers of bags need to be closed, the time-saving FT method is a good alternative PICS sealing method over the conventional twist-tie approach.

Performance of PICS bags under extreme conditions in the sahel zone of Niger.

Experiments in Niger assessed whether extreme environmental conditions including sunlight exposure affect the performance of triple-layer PICS bags in protecting cowpea grain against bruchids. Sets of PICS bags and woven polypropylene bags as controls containing 50 kg of naturally infested cowpea grain were held in the laboratory or outside with sun exposure for four and one-half months. PICS bags held either inside or outside exhibited no significant increase in insect damage and no loss in weight after 4.5 months of storage compared to the initial values. By contrast, woven bags stored inside or outside side by side with PICS bags showed several-fold increases in insects present in or on the grain and significant losses in grain weight. Grain stored inside in PICS bags showed no reduction in germination versus the initial value but there was a small but significant drop in germination of grain in PICS bags held outside (7.6%). Germination rates dropped substantially more in grain stored in woven bags inside (16.1%) and still more in woven bags stored outside (60%). PICS bags held inside and outside retained their ability to maintain internal reduced levels of oxygen and elevated levels of carbon dioxide. Exposure to extreme environmental conditions degraded the external polypropylene outer layer of the PICS triple-layer bag. Even so, the internal layers of polyethylene were more slowly degraded. The effects of exposure to sunlight, temperature and humidity variation within the sealed bags are described.

Hermetic storage of wheat and maize flour protects against red flour beetle (Tribolium castaneum Herbst).

Hermetic storage is used to protect grain against insect pests, but its utility is not limited to whole grains. We evaluated hermetically-sealed, polyethylene terephthalate (PET) bottles for preserving wheat and maize flour against red flour beetle (RFB, Tribolium castaneum, Herbst) population growth. Flours infested with RFB and kept in sealed PET bottles experienced much less weight loss over a three-month storage period than infested flour kept in unsealed bottles. RFB populations in wheat flour kept in sealed bottles did not increase, while populations in unsealed bottles grew about 50-fold during the same three-month period. Flour in sealed bottles had lower levels of oxygen and moisture than flour stored in unsealed bottles. Similar trends were observed for oxygen and moisture levels in maize flour held in hermetically sealed bottles. Hermetically-sealed bottles were effective in preventing RFB population growth and preserving maize and wheat flour. Farmers, consumers and food processors can safely store grain flour in hermetic sealed containers.

Storage of Maize in Purdue Improved Crop Storage (PICS) Bags.

Interest in using hermetic technologies as a pest management solution for stored grain has risen in recent years. One hermetic approach, Purdue Improved Crop Storage (PICS) bags, has proven successful in controlling the postharvest pests of cowpea. This success encouraged farmers to use of PICS bags for storing other crops including maize. To assess whether maize can be safely stored in PICS bags without loss of quality, we carried out laboratory studies of maize grain infested with Sitophilus zeamais (Motshulsky) and stored in PICS triple bags or in woven polypropylene bags. Over an eight month observation period, temperatures in the bags correlated with ambient temperature for all treatments. Relative humidity inside PICS bags remained constant over this period despite the large changes that occurred in the surrounding environment. Relative humidity in the woven bags followed ambient humidity closely. PICS bags containing S. zeamais-infested grain saw a significant decline in oxygen compared to the other treatments. Grain moisture content declined in woven bags, but remained high in PICS bags. Seed germination was not significantly affected over the first six months in all treatments, but declined after eight months of storage when infested grain was held in woven bags. Relative damage was low across treatments and not significantly different between treatments. Overall, maize showed no signs of deterioration in PICS bags versus the woven bags and PICS bags were superior to woven bags in terms of specific metrics of grain quality.

Grain size and grain depth restrict oxygen movement in leaky hermetic containers and contribute to protective effect.

Postharvest insect pests threaten the nutritional and financial security of smallholder farmers in the developing world. Hermetic storage, a technology that protects grain against insects by blocking their supply of oxygen, alleviates the problem of insect-caused losses. PICS (Purdue Improved Crop Storage) bags represent one hermetic technology that improves food availability and incomes of farmers. The polyethylene liners of PICS bags are sometime damaged during use, acquiring small holes or tears. Observations in the laboratory and field suggest that insect development remains localized around the point where the bag is damaged. We hypothesized that the grain within a hermetic container that has minimal localized damage (such as an insect hole), helps retard leakage of oxygen into the bag and contributes to limiting insect damage and to the overall protective effect. To test this hypothesis, we filled 4 cm dia. by 10 cm long PVC pipes with Callosobruchus maculatus (F.) infested cowpeas and sealed them with caps having a single, insect-sized hole in its center. A vertical tube positioned above the cowpea-filled PVC pipe was filled with one of three different grains (sesame, sorghum, and maize) to different depths (0, 5, 15, 30, 50 cm). Seed size and grain barrier depth significantly reduced the level of bruchid damage to the stored cowpea in the PVC container. Smaller sized grains used for the barriers retarded insect development more effectively than larger sized grains, while deeper grain depth was more effective than shallower barriers. The grain held in a hermetic container contributes in a small, but significant, way to the effectiveness of the containers.

Plant Defense Inhibitors Affect the Structures of Midgut Cells in Drosophila melanogaster and Callosobruchus maculatus.

Plants produce proteins such as protease inhibitors and lectins as defenses against herbivorous insects and pathogens. However, no systematic studies have explored the structural responses in the midguts of insects when challenged with plant defensive proteins and lectins across different species. In this study, we fed two kinds of protease inhibitors and lectins to the fruit fly Drosophila melanogaster and alpha-amylase inhibitors and lectins to the cowpea bruchid Callosobruchus maculatus. We assessed the changes in midgut cell structures by comparing them with such structures in insects receiving normal diets or subjected to food deprivation. Using light and transmission electron microscopy in both species, we observed structural changes in the midgut peritrophic matrix as well as shortened microvilli on the surfaces of midgut epithelial cells in D. melanogaster. Dietary inhibitors and lectins caused similar lesions in the epithelial cells but not much change in the peritrophic matrix in both species. We also noted structural damages in the Drosophila midgut after six hours of starvation and changes were still present after 12 hours. Our study provided the first evidence of key structural changes of midguts using a comparative approach between a dipteran and a coleopteran. Our particular observation and discussion on plant-insect interaction and dietary stress are relevant for future mode of action studies of plant defensive protein in insect physiology.

Hypoxia Treatment of Callosobruchus maculatus Females and Its Effects on Reproductive Output and Development of Progeny Following Exposure.

Modified atmospheres present a residue-free alternative to fumigants for controlling postharvest pests of grain during storage. How sub-lethal applications of this method affects the reproductive fitness of target pests, however, is still not fully understood. We examined how low levels of ambient oxygen influence the reproduction of the female cowpea bruchid (Callosobruchus maculatus), a pest of cowpea. We used three low-oxygen atmospheres-2%, 5% and 10% (v/v) oxygen-and observed their effects on: (1) the number of eggs laid by bruchids compared to insects held in normoxic (~20% oxygen) conditions; (2) the total number of eggs laid; and (3) the number of progeny that reached maturity. Low oxygen did not significantly affect the number of eggs laid during 48 or 72 h of exposure, but 2% and 5% oxygen did negatively affected total egg production. Increasing the exposure time from 48 to 72 h further depressed lifetime reproductive output. Maternal and egg exposure to hypoxia reduced the number of progeny that reached adulthood. Lower adult emergence was observed from eggs laid under low oxygen and longer exposure times. These data demonstrate that hermetic conditions depress the egg-laying behavior of cowpea bruchids and the successful development of their progeny.

RNAi for insect control: current perspective and future challenges.

The research on the RNA interference (RNAi) for the control of insect pests has made significant growth in recent years. The availability of the genomic sequences of insects has further widened the horizons for the testing of this technology to various insect groups. Different modes of application of double-stranded RNA (dsRNA) have been tested; however, the practicability of delivery of dsRNA in insects still remains the biggest challenge. Till date, the oral delivery of dsRNA in insects is one of the efficient approaches for the practical application of this technique. The uptake of dsRNA from the insect gut is mediated either by SID-1/SID-2 transmembrane proteins or by endocytosis; however, the systemic RNAi machinery still remains to be revealed in insect species. The RNAi-mediated gene knockdown has shown striking results in different insect groups, pointing it to be the upcoming technique for insect control. However, before the successful application of this technique for insect control, some potential issues need to be resolved. This review presents the account of prospects and challenges for the use of this technology for insect control.

Transcriptome sequencing, and rapid development and application of SNP markers for the legume pod borer Maruca vitrata (Lepidoptera: Crambidae).

The legume pod borer, Maruca vitrata (Lepidoptera: Crambidae), is an insect pest species of crops grown by subsistence farmers in tropical regions of Africa. We present the de novo assembly of 3729 contigs from 454- and Sanger-derived sequencing reads for midgut, salivary, and whole adult tissues of this non-model species. Functional annotation predicted that 1320 M. vitrata protein coding genes are present, of which 631 have orthologs within the Bombyx mori gene model. A homology-based analysis assigned M. vitrata genes into a group of paralogs, but these were subsequently partitioned into putative orthologs following phylogenetic analyses. Following sequence quality filtering, a total of 1542 putative single nucleotide polymorphisms (SNPs) were predicted within M. vitrata contig assemblies. Seventy one of 1078 designed molecular genetic markers were used to screen M. vitrata samples from five collection sites in West Africa. Population substructure may be present with significant implications in the insect resistance management recommendations pertaining to the release of biological control agents or transgenic cowpea that express Bacillus thuringiensis crystal toxins. Mutation data derived from transcriptome sequencing is an expeditious and economical source for genetic markers that allow evaluation of ecological differentiation.

Mitochondrial genome sequence and expression profiling for the legume pod borer Maruca vitrata (Lepidoptera: Crambidae).

We report the assembly of the 14,054 bp near complete sequencing of the mitochondrial genome of the legume pod borer (LPB), Maruca vitrata (Lepidoptera: Crambidae), which we subsequently used to estimate divergence and relationships within the lepidopteran lineage. The arrangement and orientation of the 13 protein-coding, 2 rRNA, and 19 tRNA genes sequenced was typical of insect mitochondrial DNA sequences described to date. The sequence contained a high A+T content of 80.1% and a bias for the use of codons with A or T nucleotides in the 3rd position. Transcript mapping with midgut and salivary gland ESTs for mitochondrial genome annotation showed that translation from protein-coding genes initiates and terminates at standard mitochondrial codons, except for the coxI gene, which may start from an arginine CGA codon. The genomic copy of coxII terminates at a T nucleotide, and a proposed polyadenylation mechanism for completion of the TAA stop codon was confirmed by comparisons to EST data. EST contig data further showed that mature M. vitrata mitochondrial transcripts are monocistronic, except for bicistronic transcripts for overlapping genes nd4/nd4L and nd6/cytb, and a tricistronic transcript for atp8/atp6/coxIII. This processing of polycistronic mitochondrial transcripts adheres to the tRNA punctuated cleavage mechanism, whereby mature transcripts are cleaved only at intervening tRNA gene sequences. In contrast, the tricistronic atp8/atp6/coxIII in Drosophila is present as separate atp8/atp6 and coxIII transcripts despite the lack of an intervening tRNA. Our results indicate that mitochondrial processing mechanisms vary between arthropod species, and that it is crucial to use transcriptional information to obtain full annotation of mitochondrial genomes.

Geographic distribution of phylogenetically-distinct legume pod borer, Maruca vitrata (Lepidoptera: Pyraloidea: Crambidae).

Maruca vitrata Fabricius is a pantropical lepidopteran pest of legumes. Phylogenetic analysis of a mitochondrial cytochrome c oxidase-I gene (cox1) fragment indicates that three Maruca sp. mitochondrial lineages have unique geographic distributions [lineages 1 and 2: Australia, Taiwan, and West Africa (Niger, Nigeria, and Burkina Faso), and lineage 3: Puerto Rico]. The haplotype (T30, T114) is specific to lineages 1&2 and was assayed by NsiI and SacI polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) within population samples; it was not observed in the Puerto Rican samples, but was nearly fixed among samples from West Africa, Australia and Taiwan (85.5-100%). Re-sequencing and phylogenetic analyses of PCR-RFLP defined cox1 haplotypes indicate that nucleotide diversity is highest among samples from West Africa. Phylogenetic reconstruction based upon ribosomal DNA (rDNA) internal transcribed spacer-2 (ITS-2) sequences provided additional evidence for three Maruca sp. clades. These data suggest that multiple unique Maruca species or subspecies are present worldwide, which has implications for the management of this pest species-complex.

Protease inhibitors from several classes work synergistically against Callosobruchus maculatus.

Targeting multiple digestive proteases may be more effective in insect pest control than inhibition of a single enzyme class. We therefore explored possible interactions of three antimetabolic protease inhibitors fed to cowpea bruchids in artificial diets, using a recombinant soybean cysteine protease inhibitor scN, an aspartic protease inhibitor pepstatin A, and soybean Kunitz trypsin inhibitor KI. scN and pepstatin, inhibiting major digestive cysteine and aspartic proteases, respectively, significantly prolonged the developmental time of cowpea bruchids individually. When combined, the anti-insect effect was synergistic, i.e., the toxicity of the mixture was markedly greater than that of scN or pepstatin alone. KI alone did not impact insect development even at relatively high concentrations, but its anti-insect properties became apparent when acting jointly with scN or scN plus pepstatin. Incubating KI with bruchid midgut extract showed that it was partially degraded. This instability may explain its lack of anti-insect activity. However, this proteolytic degradation was inhibited by scN and/or pepstatin. Protection of KI from proteolysis in the insect digestive tract thus could be the basis for the synergistic effect. These observations support the concept that cowpea bruchid gut proteases play a dual role; digesting protein for nutrient needs and protecting insects by inactivating dietary proteins that may otherwise be toxic. Our results also suggest that transgenic resistance strategies that involve multigene products are likely to have enhanced efficacy and durability.

Arabidopsis vegetative storage protein is an anti-insect acid phosphatase.

Indirect evidence previously suggested that Arabidopsis (Arabidopsis thaliana) vegetative storage protein (VSP) could play a role in defense against herbivorous insects. To test this hypothesis, other AtVSP-like sequences in Arabidopsis were identified through a Basic Local Alignment Search Tool search, and their transcriptional profiles were investigated. In response to methyl jasmonate application or phosphate starvation, AtVSP and AtVSP-like genes exhibited differential expression patterns, suggesting distinct roles played by each member. Arabidopsis VSP2 (AtVSP2), a gene induced by wounding, methyl jasmonate, insect feeding, and phosphate deprivation, was selected for bacterial expression and functional characterization. The recombinant protein exhibited a divalent cation-dependent phosphatase activity in the acid pH range. When incorporated into the diets of three coleopteran and dipteran insects that have acidic gut lumen, recombinant AtVSP2 significantly delayed development of the insects and increased their mortality. To further determine the biochemical basis of the anti-insect activity of the protein, the nucleophilic aspartic acid-119 residue at the conserved DXDXT signature motif was substituted by glutamic acid via site-directed mutagenesis. This single-amino acid alteration did not compromise the protein's secondary or tertiary structure, but resulted in complete loss of its acid phosphatase activity as well as its anti-insect activity. Collectively, we conclude that AtVSP2 is an anti-insect protein and that its defense function is correlated with its acid phosphatase activity.

"Active" refuges can inhibit the evolution of resistance in insects towards transgenic insect-resistant plants.

Negative cross-resistance (NCR) toxins that hitherto have not been thought to have practical uses may indeed be useful in the management of resistance alleles. Practical applications of NCR for pest management have been limited (i) by the scarcity of high toxicity NCR toxins among pesticides, (ii) by the lack of systematic methodologies to discover and develop such toxins, as well as (iii) by the lack of deployment tactics that would make NCR attractive. Here we present the concept that NCR toxins can improve the effectiveness of refuges in delaying the evolution of resistance by herbivorous insect pests to transgenic host plants containing insecticidal toxins. In our concept, NCR toxins are deployed in the refuge, and thus are physically separated from the transgenic plants containing the primary plant-protectant gene (PPPG) encoding an insecticidal toxin. Our models show: (i) that use of NCR toxins in the refuge dramatically delays the increase in the frequency of resistance alleles in the insect population; and (ii) that NCR toxins that are only moderately effective in killing insects resistant to the PPPG can greatly improve the durability of transgenic insecticidal toxins. Moderately toxic NCR toxins are more effective in minimizing resistance development in the field when they are deployed in the refuge than when they are pyramided with the PPPG. We explore the potential strengths and weaknesses of deploying NCR toxins in refuges.

Soyacystatin N inhibits proteolysis of wheat alpha-amylase inhibitor and potentiates toxicity against cowpea weevil.

Genetic engineering may be used to introduce multiple insect resistance genes with different modes of action into crop plants. We explored the possible interactions of two differing gene products fed in the diet of cowpea weevil, Callosobruchus maculates (F.), a stored grain pest. The soybean cysteine protease inhibitor soyacystatin N (scN) and alpha-amylase inhibitor (alphaAI) from wheat have defensive function against this coleopteran. When artificial seeds containing both scN and alpha(AI) were infested with eggs of C. maculatus, the delays in larval development were longer than was predicted by summing the developmental delays seen when larvae were fed a diet containing the individual proteins, indicating that the effects of scN and alpha(AI) are synergistic. Alpha(AI) was readily hydrolyzed when incubated with insect gut extract. This proteolytic degradation was inhibited by scN, but not by Kunitz inhibitor (a serine protease inhibitor). Thus, degradation of alpha(AI) was due to proteolysis by insect digestive cysteine proteases. These data suggest that C. maculatus uses digestive enzymes not only to function in food protein digestion but also to defend the insects themselves by helping reduce the concentration of a toxic dietary protein.