PICS bags safely store unshelled and shelled groundnuts in Niger.

We conducted an experiment in Niger to evaluate the performance of hermetic triple layer (Purdue Improved Crop Storage- PICS) bags for the preservation of shelled and unshelled groundnut Arachis hypogaea L. Naturally-infested groundnut was stored in PICS bags and woven bags for 6.7 months. After storage, the average oxygen level in the PICS bags fell from 21% to 18% (v/v) and 21%-15% (v/v) for unshelled and shelled groundnut, respectively. Identified pests present in the stored groundnuts were Tribolium castaneum (Herbst), Corcyra cephalonica (Stainton) and Cryptolestes ferrugineus (Stephens). After 6.7 months of storage, in the woven bag, there was a large increase in the pest population accompanied by a weight loss of 8.2% for unshelled groundnuts and 28.7% for shelled groundnut. In PICS bags for both shelled and unshelled groundnuts, by contrast, the density of insect pests did not increase, there was no weight loss, and the germination rate was the same compared to that recorded at the beginning of the experiment. Storing shelled groundnuts in PICS bags is the most cost-effective way as it increases the quantity of grain stored.

Quality of maize for sale in markets in Benin and Niger.

A follow-up study on the quality of maize for sale in West African public markets was carried out in Benin and Niger from August 15-28, 2013. Complementing the earlier study, this present assessment included not only retailers but also wholesalers and maize producers. Samples were evaluated for parameters related to the physical quality of the maize and for aflatoxin contamination. Most maize value chain actors process their offered grain using traditional methods for threshing, winnowing and drying. Maize for sale in the markets surveyed had an average moisture content ranging between 12 and 14%. Non-grain impurities amounted to 0-2.3% while mouldy grains ranged between 0.2 and 0.8%. The impurity level in grain was three times higher among wholesalers compared to retailers and producers. An insect pest, the Larger Grain Borer (Prostephanus truncatus (Horn) was found only in Benin but Sitophilus zeamais Motschulsky, Cryptolestes ferrugineus Stephens, and Tribolium castaneum Herbst, were present in maize for sale in the markets in of both countries. Insect pest frequency was 16 times higher in wholesalers' grain compared to that of retailers and producers. Aflatoxin levels exceeding the accepted standard of 20 ppb were noted in markets in both countries. The highest proportion of aflatoxin-contaminated maize was in wholesalers' grain in Malanville market.

Triple bag hermetic technology for controlling a bruchid (Spermophagus sp.) (Coleoptera, Chrysomelidae) in stored Hibiscus sabdariffa grain.

We assessed the performance of hermetic triple layer Purdue Improved Crop Storage (PICS) bags for protecting Hibiscus sabdariffa grain against storage insects. The major storage pest in the grain was a bruchid, Spermophagus sp.. When we stored infested H. sabdariffa grain for six months in the woven polypropylene bags typically used by farmers, the Spermophagus population increased 33-fold over that initially present. The mean number of emergence holes per 100 seeds increased from 3.3 holes to 35.4 holes during this time period, while grain held for the same length of time in PICS bags experienced no increase in the numbers of holes. Grain weight loss in the woven control bags was 8.6% while no weight loss was observed in the PICS bags. Seed germination rates of grain held in woven bags for six months dropped significantly while germination of grain held in PICS bags did not change from the initial value. PICS bags can be used to safely store Hibiscus grain after harvest to protect against a major insect pest.

Triple-Layer Plastic Bags Protect Dry Common Beans (Phaseolus vulgaris) Against Damage by Acanthoscelides obtectus (Coleoptera: Chrysomelidae) During Storage.

Fumigated dry common beans (Phaseolus vulgaris L.) that were artificially infested with Acanthoscelides obtectus Say, and others that were not artificially infested, were stored in hermetic triple-layer PICS (Lela Agro, Kano, Nigeria) or woven polypropylene (PP) bags for 6 mo at ambient laboratory temperature conditions of 22.6 ± 1.9°C and 60.1 ± 4.3% relative humidity. In an additional trial, beans contained in PP bags were treated with Actellic Super dust before introducing A. obtectus. Moisture content, number of live adult A. obtectus, seed damage, weight loss, and seed germination were determined at monthly intervals. At 6 mo, beans stored in PICS bags retained higher moisture than those stored in PP bags, but in all treatments the moisture level remained below that recommended for safe storage of beans. In the PICS bags, proliferation of A. obtectus did not proceed and at 6 mo, beans stored in these bags did not have insect-inflicted seed damage or weight loss. In contrast, seed damage and weight loss in PP bags exceeded economic threshold after 1 mo in the absence of Actellic Super dust (Syngenta Crop protection AG, Basle, Switzerland), and after 2 mo in the presence of it. Germination of beans stored in PP bags decreased greatly whereas the beans stored in PICS bags did not show reduced germination. Chemical free storage of common beans in PICS bags protects them against damage by A. obtectus.

The spatial genetic differentiation of the legume pod borer, Maruca vitrata F. (Lepidoptera: Crambidae) populations in West Africa.

The legume pod borer, Maruca vitrata, is an endemic insect pest that causes significant yield loss to the cowpea crop in West Africa. The application of population genetic tools is important in the management of insect pests but such data on M. vitrata is lacking. We applied a set of six microsatellite markers to assess the population structure of M. vitrata collected at five sites from Burkina Faso, Niger and Nigeria. Observed polymorphisms ranged from one (marker 3393) to eight (marker 32008) alleles per locus. Observed and expected heterozygosities ranged from 0.0 to 0.8 and 0.0 to 0.6, respectively. Three of the loci in samples from Nigeria and Burkina Faso deviated significantly from Hardy-Weinberg Equilibrium (HWE), whereas no loci deviated significantly in samples from Niger. Analysis of molecular variance (AMOVA) indicated that 67.3% level of the genetic variation was within individuals compared to 17.3% among populations. A global estimate of F ST=0.1 (ENA corrected F ST=0.1) was significant (P⩽0.05) and corroborated by pairwise F ST values that were significant among all possible comparisons. A significant correlation was predicted between genetic divergence and geographic distance between subpopulations (R2=0.6, P=0.04), and cluster analysis by the program STRUCTURE predicted that co-ancestry of genotypes were indicative of three distinct populations. The spatial genetic variance among M. vitrata in West Africa may be due to limited gene flow, south-north seasonal movement pattern or other reproductive barriers. This information is important for the cultural, chemical and biological control strategies for managing M. vitrata.

Bowman-Birk inhibitor affects pathways associated with energy metabolism in Drosophila melanogaster.

Bowman-Birk inhibitor (BBI) is toxic when fed to certain insects, including the fruit fly, Drosophila melanogaster. Dietary BBI has been demonstrated to slow growth and increase insect mortality by inhibiting the digestive enzymes trypsin and chymotrypsin, resulting in a reduced supply of amino acids. In mammals, BBI influences cellular energy metabolism. Therefore, we tested the hypothesis that dietary BBI affects energy-associated pathways in the D. melanogaster midgut. Through microarray and metabolomic analyses, we show that dietary BBI affects energy utilization pathways in the midgut cells of D. melanogaster. In addition, ultrastructure studies indicate that microvilli are significantly shortened in BBI-fed larvae. These data provide further insights into the complex cellular response of insects to dietary protease inhibitors.

Transcriptional signatures in response to wheat germ agglutinin and starvation in Drosophila melanogaster larval midgut.

One function of plant lectins such as wheat germ agglutinin is to serve as defences against herbivorous insects. The midgut is one critical site affected by dietary lectins. We observed marked cellular, structural and gene expression changes in the midguts of Drosophila melanogaster third instar larvae that were fed wheat germ agglutinin. Some of these changes were similar to those observed in the midguts of starved D. melanogaster. Dietary wheat germ agglutinin caused shortening, branching, swelling, distortion and in some cases disintegration of the midgut microvilli. Starvation was accompanied primarily by shortening of the microvilli. Microarray analyses revealed that dietary wheat germ agglutinin evoked differential expression of 61 transcripts; seven of these were also differentially expressed in starved D. melanogaster. The differentially transcribed gene clusters in wheat germ agglutinin-fed larvae were associated with (1) cytoskeleton organization; (2) digestive enzymes; (3) detoxification reactions; and (4) energy metabolism. Four possible transcription factor binding motifs were associated with the differentially expressed genes. One of these exhibited substantial similarity to MyoD, a transcription factor binding motif associated with cellular structures in mammals. These results are consistent with the hypothesis that wheat germ agglutinin caused a starvation-like effect and structural changes of midgut cells of D. melanogaster third-instar larvae.

Cowpea bruchid midgut transcriptome response to a soybean cystatin--costs and benefits of counter-defence.

The insect digestive system is the first line of defence protecting cells and tissues of the body from a broad spectrum of toxins and antinutritional factors in its food. To gain insight into the nature and breadth of genes involved in adaptation to dietary challenge, a collection of 20 352 cDNAs was prepared from the midgut tissue of cowpea bruchid larvae (Callosobruchus maculatus) fed on regular diet and diets containing antinutritional compounds. Transcript responses of the larvae to dietary soybean cystatin (scN) were analysed using cDNA microarrays, followed by quantitative real-time PCR (RT-PCR) confirmation with selected genes. The midgut transcript profile of insects fed a sustained sublethal scN dose over the larval life was compared with that of insects treated with an acute high dose of scN for 24 h. A total of 1756 scN-responsive cDNAs was sequenced; these clustered into 967 contigs, of which 653 were singletons. Many contigs (451) did not show homology with known genes, or had homology only with genes of unknown function in a Blast search. The identified differentially regulated sequences encoded proteins presumptively involved in metabolism, structure, development, signalling, defence and stress response. Expression patterns of some scN-responsive genes were consistent in each larval stage, whereas others exhibited developmental stage-specificity. Acute (24 h), high level exposure to dietary scN caused altered expression of a set of genes partially overlapping with the transcript profile seen under chronic lower level exposure. Protein and carbohydrate hydrolases were generally up-regulated by scN whereas structural, defence and stress-related genes were largely down-regulated. These results show that insects actively mobilize genomic resources in the alimentary tract to mitigate the impact of a digestive protease inhibitor. The enhanced or restored digestibility that may result is possibly crucial for insect survival, yet may be bought at the cost of weakened response to other stresses.

Morphometry and ultrastructure of prismatic cristae in mitochondria of a crayfish muscle. A hypothesis of the structural principle.

Approximately 40% of the mitochondria in the sphincter muscle of the crayfish vas deferens have prismatic-type cristae. In cross section, the angular cristae have either triangular or rhomboid profiles which are surrounded by a hexagonal array of electron-dense dots. In longitudinal section, these mitochondria exhibit both thick and thin parallel lines, which represent cristae and filaments, respectively. It is postulated that the matrix of the prismatic-type mitochondria is packed with rodlets composed of an electron-dense core and a less dense shell. Close packing of these rodlets results in the regular hexagonal dot array. Deletion of fascicles of 3 or 4 rodlets results in spaces with triangular or rectangular cross sections. Lining of these spaces with membranes results in cristae with triangular or rhomboid cross sections.

Formamidine pesticides: octopamine-like actions in a firefly.

The formamidine pesticide chlordimeform and its N-demethylated metabolites cause the light organ of the firefly Photinus pyralis L. to glow brightly. Monodemethyl chlordimeform is active at doses as low as 5 nanograms per insect when applied topically. This action is postsynaptic and probably involves membrane-bound receptors since cyproheptadine blocks the glows induced by both monodemethyl chlordimeform and octopamine, the putative neurotransmitter in the light organ. The pesticidal and pestistatic properties of the formamidines may result from actions on octopaminergic systems.

Efficacy of Hypoxia Against Tribolium castaneum (Coleoptera: Tenebrionidae) Throughout Ontogeny.

Hermetic grain storage technology offers a viable chemical-free approach to control storage insects. However, there is limited knowledge on how hypoxia affects the survival of insect life stages during grain storage in hermetic bags. We exposed Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) eggs (2 d), young larvae (7 d), old larvae (21 d), pupae (28 d), and adults (2 d after emergence) to 2, 4, 8, and 20.9% oxygen levels for 1, 3, 5, 10, and 15 d and assessed subsequent mortality. At 2% oxygen, complete mortality was achieved in 3 d for eggs and young larvae, 10 d for old larvae and pupae, and 15 d for adults. At 4% oxygen, 15 d were required to kill all eggs and old larvae but not the other insect life stages. At 8% oxygen after 15 d, complete mortality of any insect life stage was not observed; but even a relatively short exposure (1-3 d) caused significant developmental delays in immature insects. Our study shows potential utility of hermetic technology for control of T. castaneum, but internal oxygen should be maintained below 2% level for at least 15 d for complete control. Increased oxygen levels improved the development of all insect life stages leading to increased adult emergence. There is a need to explore exposure time required to achieve complete mortality of all insect life stage above the 2% oxygen level.

Transcriptomic profiles of Drosophila melanogaster third instar larval midgut and responses to oxidative stress.

Oligoarray analysis was used to determine the number and nature of genes expressed in third instar Drosophila melanogaster larval midguts. The majority of transcripts were associated with protein synthesis and metabolism. Serine proteases were the main proteolytic enzymes detected. Some 40% of the cytochrome P450 genes and 74% of the glutathione S transferases (GSTs) in the genome of D. melanogaster were observed to be expressed in the midgut by oligoarray analysis. We also identified potential transcription factor binding motifs (TFBMs) of P450s, GSTs and carboxylesterases. Many of the midgut-expressed GST genes contained candidate TFBMs homologous to TFBMs in mammals that have been associated with responses to oxidative stress. We also investigated the response of GSTs in the midgut to dietary H2O2, which showed a dosage-based differential response.

Identification of N-acetylglucosamine binding residues in Griffonia simplicifolia lectin II.

Primary structure and crystallographic data of several legume lectins were used to predict the involvement in carbohydrate binding of six amino acid residues (Asp88, Glu108, Tyr134, Asn136, Leu226 and Gln227) in Griffonia simplicifolia lectin II (GS-II). The functional involvement of these residues was evaluated by assessing GlcNAc binding of modified forms of GS-II in which these residues were eliminated in truncated peptides or systematically substituted with other amino acids by site-specific mutations. Mutations at Asp88, Tyr134 or Asn136 eliminated GlcNAc binding activity by GS-II, while those at Glu108, Leu226 or Gln227 did not alter the activity. The former three amino acids were functionally essential for carbohydrate binding by GS-II presumably through hydrogen bonding to and hydrophobic interactions with GlcNAc. Although an Asp or Gly substitution for Tyr134 eliminated GlcNAc affinity, substitution with Phe did not appreciably affect binding. Despite the fact that mutations to Leu226 and Gln227 did not alter carbohydrate binding, a truncated form of GS-II lacking these residues no longer exhibited carbohydrate binding affinity.

A plant defensive cystatin (soyacystatin) targets cathepsin L-like digestive cysteine proteinases (DvCALs) in the larval midgut of western corn rootworm (Diabrotica virgifera virgifera).

Feeding bioassay results established that the soybean cysteine proteinase inhibitor N (soyacystatin N, scN) substantially inhibits growth and development of western corn rootworm (WCR), by attenuating digestive proteolysis [Zhao, Y. et al. (1996) Plant Physiol. 111, 1299-1306]. Recombinant scN was more inhibitory than the potent and broad specificity cysteine proteinase inhibitor E-64. WCR digestive proteolytic activity was separated by mildly denaturing SDS-PAGE into two fractions and in-gel assays confirmed that the proteinase activities of each were largely scN-sensitive. Since binding affinity to the target proteinase [Koiwa, H. et al. (1998) Plant J. 14, 371-380] governs the effectiveness of scN as a proteinase inhibitor and an insecticide, five peptides (28-33 kDa) were isolated from WCR gut extracts by scN affinity chromatographic separation. Analysis of the N-terminal sequence of these peptides revealed similarity to a cathepsin L-like cysteine proteinase (DvCAL1, Diabrotica virgifera virgifera cathepsin L) encoded by a WCR cDNA. Our results indicate that cathepsin L orthologs are pivotal digestive proteinases of WCR larvae, and are targets of plant defensive cystatins (phytocystatins), like scN.

Central projections of axons from taste hairs on the labellum and tarsi of the blowfly, Phormia regina Meigen.

Taste hairs are located on the labellum and tarsi of blowflies. These multimodal hairs consist of four functionally distinct chemoreceptors and a mechanoreceptor. By staining selected multimodal hairs, we sought to identify the central projection patterns of multiple and single axons from those hairs. On each side of the labellum there are 11 "largest" hairs (LH). The neurons associated with the anteriormost (LH-1), posteriormost (LH-11), and one lateral (LH-6) hair on the labellum were stained selectively with cobaltous sulfide. The overall projection pattern in the central nervous system (CNS) for axons from LH-1 and LH-11 is similar and differs markedly from axons from LH-6. At least three individual axon-projection patterns were determined for each labellar hair filled, indicating a partial functional organization for axons from multimodal hairs. One identified axon, the dorsalmost axon, has terminal arborizations that do not differ with the location of its associated hair. Another axon, thicker than the others, projects to a region that is distinct from the four thin axons. Within this region the arborizations of the thick axons occupy different areas depending on the location of their associated hair. Neurons from the largest hairs on the distalmost tarsomere (D5) of each leg were also stained and consisted of one thick and four thin axons. All axons except one thin axon from tarsal D5 hairs terminate in their respective leg neuromeres. The remaining thin axon projects to the suboesophageal ganglion ipsilateral to the hair filled and terminates in the same region as a branch of the labellar dorsalmost axon. These data suggest that axonal arbors from multimodal hairs have a limited functional and somatotopic organization in the blowfly CNS.

Two-dimensional Protein Patterns in Labronema, Aporcelaimellus, and Eudorylaimus (Nematoda: Dorylaimida).

Two-dimensional polyacrylamide gel electrophoretic patterns of proteins for two isolates of Labronema from Indiana were nearly identical to the pattern for L. vulvapapillatum from Europe. The pattern for a nominal isolate of L. pacificum from Florida was very different from the patterns of nominal L. pacificum isolates from Hawaii and Fiji (which had patterns very similar to each other). Patterns for four other isolates (in Eudorylaimus and Aporcelaimellus) were different from the Labronema patterns and from each other, although some constellations of protein spots were shared among all the isolates. The study demonstrates the utility of 2-D PAGE for clarifying taxonomic problems that cannot be resolved using classical morphological data alone.

Heterodera glycines in Indiana: III. 2-D Protein Patterns of Geographical Isolates.

Protein patterns obtained by two-dimensional polyacrylamide gel electrophoresis for three isolates of Heterodera glycines from southern Indiana appear qualitatively similar and have higher pairwise Jaccard similarity coefficients with each other than with isolates from northern Indiana. Three isolates from three northern counties share proteins not present in the southern isolates, but as a group the northern isolates are less similar to each other than are the southern Indiana isolates.

Two-Dimensional Protein Patterns in Heterodera glycines.

Two-dimensional polyacrylamide gel electrophoretic protein patterns of H. glycines from southern Indiana (Posey County) and northern Indiana (Pulaski County) were largely similar, but many differences existed. The pattern of the Posey isolate was similar to patterns from isolates collected in other areas of the United States. Unique dense protein spots in the pattern of an isolate from Hokkaido, Japan, distinguished it from patterns of six U.S. isolates.