Effect of seed lectins from Phaseolus vulgaris on the development of larvae of Callosobruchus maculatus; mechanism of toxicity

Seeds of the kidney bean (Phaseolus vulgaris) are toxic to developing larvae of the bruchid beetle (Callosobruchus maculatus), a major storage pest of many legumes. Insect feeding trials were carried out whereby the albumin and globulin protein fractions from seeds of P. vulgaris were incorporated into artificial seeds. Both fractions were shown to be toxic and to contain haemagglutinating activity, implicating the seed lectins as being involved in seed resistance. Further feeding trials using different P. vulgaris lectin preparations confirmed the toxicity of these lectins and suggested that it was the E-type lectin subunits (erythrocyte-binding) which were the major antimetabolites. Indirect immunofluorescence investigations using monospecific antisera for globulin lectins showed that the lectins, when ingested by the larvae, bound to the midgut epithelial cells. It was suggested that the mechanism of lectin toxicity in this instance is analogous to that known to occur in the rat, namely that the ingested lectin causes disruption of the epithelial cells of the larval midgut leading to breakdown of the transport of nutrients into these cells, and the absorption of potentially harmful substances. This is the first time that evidence for the mechanism of lectin toxicity has been obtained in insects.     

Deterrent and insecticidal properties of bean seed (Phaseolus vulgaris L.) whole meal or protein extract incorporated into the diet of Callosobruchus maculatus (F.) (Coleoptera: Bruchidae)

Callosobruchus maculatus, a pest that causes serious damage to chickpea Cicer arietinum, cannot develop in the seeds of Phaseolus or Vigna spp. which contain lectins. The insecticidal activity towards C. maculatus in these seeds is attributed both to lectins with specific affinity to N-acetylglucosamine, the major component of insect chitin, and to alpha-amylase inhibitors (lectin-like proteins). The insecticidal properties of bean meal or bean protein extracts from different sources towards different pest species are variable and need to be experimentally evaluated. The main objective of this study was to determine through a feeding trial on artificial chickpea seed the potential of bean seed meal from a wild bean Vigna caracalla, four varieties of Phaseolus vulgaris, and of a protein extract of P. vulgaris seed, to alter different life history traits of C. maculatus. The chickpea weevil was set up on artificial chickpea seeds containing different amounts of bean meal to observe the effects on female oviposition, percentage of development to adulthood and juvenile development time. These traits were combined in a composite index measuring the alteration of the multiplication rate of C. maculatus fed on artificial seed. The activity of lectin-like extracts was observed on chickpea artificial seed spiked with bean seed extract. Incorporation of bean flour at a rate of 10 and 20% in chickpea artificial seed significantly decreased C. maculatus female fecundity, percentage of adult emergence, and greatly increased the development time. Feeding trials with protein extracts of P. vulgaris reduced fecundity and survival of C. maculatus. Incorporation of 10% V. caracalla bean seed meal in chickpea artificial seed, reduced the multiplication potential of C. maculatus by over 90% showing that bean seed lectin extracts are worthy of further investigation for post-harvest infestation control.     

Mechanism of seed lectin tolerance by a major insect storage pest of Phaseolus vulgaris,acanthoscelides obtectus

The bean weevil Acanthoscelides obtectus (Say) is a major storage peat of Phaseolus vulgaris L (kidney, haricot bean). The seeds of P vulgaris contain high levels (up to 30 mg g^?1 DM) of lectin, which has been shown to be toxic towards larvae of the related bruchid storage pest of cowpea, Callosobruchus maculatus F. The lack of toxicity of this lectin towards larvae of A obtectus is demonstrated. Unlike the strong binding of lectin to the midgut epithelium observed in larvae of C maculatus, no binding of lectin molecules was found to occur in the gut epithelial cells of A obtectus. This observation provides the basis for a hypothesis explaining the lack of toxicity of P vulgaris lectin towards A obtectus. Assays of proteolytic activity in gut homogenates of C maculatus and A obtectus suggest that the difference in susceptibility of the two insects towards the toxic effects of the lectin is not due to differential inactivation by proteolysis. Besides its effects on larval development, the lectin has a further effect at pupation, causing disruption of adipose tissues in C maculatus but having no effect on A obtectus.     

Identification,biochemical characterization and biological role of defense proteins from common bean genotypes seeds in response to Callosobruchus maculatus infestation

Common bean is a legume of significant socioeconomic importance and is cultivated worldwide. This crop is affected by several pests and diseases, which cause considerable economic losses and reduce yield. In recent years, several studies have demonstrated the role of proteins and peptides with activity against a wide range of insects and pathogens. The objective of this work was to identify defense proteins, such as antimicrobial peptides, protease and amylases inhibitors in common bean genotypes and evaluate the relationship of these proteins with Phaseolus vulgaris seed resistance to Callosobruchus maculatus infestation. Nineteen common bean genotypes were subjected to protein extraction, pH 5.4, and precipitation with ammonium sulfate at 70% saturation. The obtained extracts were separated by tricine gel electrophoresis. Experiments were carried out with natural seeds of common beans and artificial seeds (Vigna unguiculata seeds covered with seed coats of common beans) to evaluate the rate of oviposition and development of the insect species Callosobruchus maculatus. Lipid-transfer proteins were identified in nine genotypes whereas defensins were present in all genotypes. The inhibitory activity of α-amylases and trypsin and fungal development were determined in crude extracts (50 μg mL⁻¹). The results also indicated that the extracts from all bean genotypes inhibited the activity of human salivary α-amylase and C. maculatus larval α-amylase. Except for the extracts of four genotypes, all other extracts inhibited trypsin activity. None of the extracts from the evaluated bean genotypes inhibited the growth of tested fungi. Natural seeds from all genotypes did not inhibit insect oviposition, however, the larvae did not survive after feeding on these seeds. Artificial seeds containing seed coat flour of all genotypes inhibited the oviposition of C. maculatus, indicating that the seed coat was also repellent to insect.     

Biochemical basis of insect resistance in Vigna unguiculata

The bruchid beetle Callosobruchus maculatus (F.) causes extensive damage to seeds of the cowpea, Vigna unguiculata (L.) (Walp.), when this important tropical foodstuff is stored. A variety of cowpea resistant to attack by this pest has been described. In the present work seeds of a number of cowpea varieties, including the resistant one, were tested for the presence of a physical resistance to C. maculatus, in terms of repulsion of oviposition or of failure of larvae to enter the seeds. No evidence to suggest the presence of a physical resistance was found. When seeds of cowpea varieties were tested for the presence of various antimetabolic secondary compounds, only inhibitory activity against trypsin and, to a much lesser extent, chymotrypsin, could be detected. The resistant variety of cowpea contained a significantly higher level of inhibitors, about twice as much as any other variety. A proteinase inhibitor active against trypsin was purified from cowpea varieties by affinity chromatography on trypsin-Sepharose. The purified inhibitor was shown to inhibit chyraotrypsin also, in such proportions as to account for chymotrypsin inhibition by seed extracts. The inhibitor was shown to consist of a number of isoinhibitors by gel electrophoresis and isoelectric focusing, but no qualitative differences in the inhibitor between varieties could be detected. The antimetabolic nature of the cowpea trypsin inhibitor was confirmed by insect feeding trials in which various protein fractions were added to a basic meal and the effect on larval survival noted. The albumin proteins of cowpea (containing the trypsin inhibitors) at a level of 10% were toxic to larvae of C. maculatus whereas the globulin fractions were not. Further, if cowpea trypsin inhibitor was removed from the albumin proteins they ceased to be toxic. When purified cowpea trypsin inhibitor was added to the basic meal it was shown that a level slightly less than that found in the resistant variety of cowpea caused complete mortality of larvae, whereas lower levels had lesser or no effect. It is concluded that this example of insect resistance in the cowpea is due to an elevated level of trypsin inhibitor.     

Characterization of new sources of mungbean (Vigna radiata (L.) Wilczek) resistance to bruchids, Callosobruchus spp. (Coleoptera: Bruchidae)

The bruchid beetles Callosobruchus chinensis (L.) and C. maculatus (F.) are destructive pests of stored mungbean (Vigna radiata (L.) Wilczek). The development of resistant mungbean varieties to manage bruchids is a major breeding objective. In the present study, we investigated the characteristics of resistance to C. chinensis and C. maculatus in two new resistant mungbean accessions V1128 and V2817, and two previously reported resistant accessions V2709 and V2802, compared to the susceptible mungbean cultivar, KPS1. Evaluation for resistance to both bruchid species using whole and decorticated seeds revealed that V1128 and V2817 were free from damage, V2709 and V2802 showed partial damage with low or moderate number of bruchids emerging from seeds, and KPS1 showed complete damage with the highest number of bruchids emerged. Comparison of the seeds harvested at seed filling, early maturity and full maturity stages revealed that the percentage of damaged seeds from resistant accessions was lower at all stages compared with KPS1. V1128 and V2817 showed complete resistance against both bruchids regardless of when their seeds were harvested, while resistance in V2709 and V2802 were most pronounced at full maturity, and KPS1 was totally damaged at all times tested. These results suggest that the chemical factor(s) conferring resistance is synthesized as early as the seed filling stage. Evaluation of resistance using artificial seeds showed that increasing the percentage of resistant seed powder adversely affected bruchid growth and development. The number of adults emerging from seeds and number of damaged seeds decreased while adult developmental period increased as the proportion of resistant seed powder increased. The weight of emerging male and female adults of C. maculatus was lighter than those from the seeds containing susceptible seed powder alone. However, C. chinensis adults were not affected by the same test. The results suggest that biochemical(s) in cotyledon tissue are responsible for the resistance and the seed coat had no protective role against the bruchids. Although all four resistant accessions evaluated are useful for mungbean breeding, V1128 and V2817 show complete resistance to both C. chinensis and C. maculatus. Thus, these two new resistant sources may be the most effective for breeding purposes.     

Legume seed vicilins (7S storage proteins) interfere with the development of the cowpea weevil (Callosobruchus maculatus (F))

Vicilins (7S storage proteins) isolated from the seeds of the legumes Vigna unguiculata (cowpea), Vigna angularis (adzuki bean), Canavalia ensiformis (jack bean), Glycine max (soybean), Phaseolus vulgaris (common bean) and Phaseolus lunatus (lima bean) were shown to be immunologically related and to bind to a chitin matrix. The effect of the isolated vicilins on the development of the cowpea weevil Callosobruchus maculatus was examined. Vicilins from all non-host seeds, including those of the C maculatus-resistant cowpea line IT81D-1045, strongly inhibited larval development (ED₅₀ of 1·07±0·32% to 2·22±0·64%). Vicilins from the C maculatus-susceptible cowpea CE-31 and adzuki bean seeds were the exception with ED₅₀ of 6·25±0·75% and 5·40±1·54%, respectively. These results correlate well with the host range of C maculatus and are possibly a reflection of the low digestibility of vicilins by insect midgut proteinases in addition to the ability they show in binding to chitin-containing structures like the ones found in the bruchid midguts. © 1998 SCI.     

Analysis of the insecticidal activity of methylisothiocyanate on Callosobruchus maculatus (F.) (Coleoptera: Bruchidae) and its parasitoid Dinarmus basalis (Rondani) (Hymenoptera: Pteromalidae)

The bruchid Callosobruchus maculatus causes major losses during the storage of the seeds of Vigna unguiculata in West Africa. An endemic parasitoid, the pteromalid Dinarmus basalis is present in the stores and can reduce the increase in bruchid populations. African farmers often place in the stores the leaves of a shrub, Boscia senegalensis Lam.(Capparaceae), which release methylisothiocyanate (MITC). This compound is toxic to adult bruchids and could reduce seed losses but its influence on the bruchid natural enemy D. basalis was hitherto unknown. The susceptibility of C. maculatus and D. basalis to MITC was assessed under laboratory conditions. The susceptibility of adults of both species towards MITC were similar. After a 24 h exposure, the LC₅₀ was respectively 0.48 and 0.54 mg/l for C. maculatus and D. basalis. The ovicidal activity of this compound was high; the LC₅₀ of C. maculatus eggs was 0.04 mg/l after a 24 h exposure to MITC. High MITC concentrations only slightly affected the survival of C. maculatus during its post-embryonic development in the seeds of V. unguiculata. Gas chromatographic analysis demonstrated that 25-35% of the MITC present in the atmosphere of the experimental jars was absorbed by the seeds but concentrations inside the cotyledons were too low to influence the survival of the C. maculatus larvae. The D. basalis larvae developing at the expense of their host inside the larval galleries, were more affected by the treatment. The introduction of B. senegalensis releasing MITC in the storage systems could reduce the density of the parasitoid population and so increase the seed losses by permitting the development of the bruchid population.     

Correlation between biophysical seed characteristics of rice bean,Vigna umbellata (Fabaceae: Faboideae: Phaseoleae) and the development of Callosobruchus maculatus (Coleoptera: Chrysomelidae: Bruchinae)

The bruchid, Callosobruchus maculatus is a serious pest of stored pulses. Four biophysical seed characteristics viz. Seed size, texture of seed surface, thickness of seed coat and seed hardness of 10 genotypes of rice bean, a wild pulse crop were investigated for assessing mechanism of resistance against C. maculatus. Oviposition preference and larval penetration in seed coat did not differ significantly among different test genotypes. Highest number of larvae which developed up to last larval/pupal instar were observed in susceptible check variety of mungbean, PAU 911 (30.00) and lowest in rice bean genotype, LRB 535 (0.83). Highest and lowest seed weight was noticed in rice bean genotype, LRB 529 (6.81 g) and PAU 911 (3.88 g), respectively. Almost similar seed coat thickness was recorded among different test genotypes. Maximum seed hardness was noted in genotype LRB 507 (6.45 kg) whereas minimum in PAU 911 (2.00 kg) among all test genotypes. Any remarkable correlation of biophysical seed characteristics of test genotypes with ovipositon, larval penetration and development of C. maculatus was not observed. Thus, biophysical seed characteristics of rice bean did not suggest to play a vital role in imparting resistance against C. maculatus.     

The effects of α-amylase inhibitors on insect storage pests: Inhibition of α-amylase in vitro and effects on development in vivo

Protein α-amylase inhibitors were prepared from wheat and their effects tested against insect storage pests both in vitro against the insect α-amylases and in vivo in insect feeding trials. Inhibitor fraction A was found to inhibit porcine pancreatic α-amylase but not insect α-amylases, whereas fractions B, C and D (0.28) did not inhibit porcine pancreatic α-amylase but were strong inhibitors of digestive α-amylases from larvae of Tribolium confusum, a storage pest of wheat products, and Callosobruchus maculatus, a storage pest of legume seeds. Fraction D, which was a single polypeptide of M_r 13 000 was the most effective inhibitor in vitro. It would appear that the degree of inhibition by the wheat α-amylase inhibitor preparations can be correlated with the presence of the M_r 13 000 (0.28) polypeptide since the purer this polypeptide the stronger was the inhibition; fraction A which contained two polypeptides of M_r 60 000 and 58 000 caused no inhibition. The effects of fractions B and C on larval development were determined in insect feeding trials. With C. maculatus both fractions were toxic, their relative effectiveness being directly paralleled by their effectiveness observed in vitro. Only fraction C was tested against T. confusum in feeding trials. Despite this fraction being equally effective against both pests in vitro it had very little effect upon larval development of T. confusum in vivo, thus suggesting that this organism is able to detoxify the wheat α-amylase inhibitors. As far as the authors are aware, this is the first time that the effects of identified inhibitor fractions have been monitored both in vitro and in vivo. The results, in contrast to previous proposals, suggest that selecting wheat varieties for high α-amylase inhibitory activity may not be a very reliable criterion in selecting for insect resistance.     

Impregnated bags for safer storage of legume grains in West and Central Africa

The use of jute bags impregnated with aqueous extracts from two insecticidal plants, Chenopodium ambrosioides and Lantana camara, to reduce damage to stored legume seeds by two major bruchid insect pests, Acanthoscelides obtectus and Callosobruchus maculatus, was compared with a direct seed-treatment method using plant powders. In the first method bags were treated with a 10% concentration of the aqueous plant extract and filled with bean or cowpea seeds, whereas in the second method seeds were thoroughly mixed with leaf powder from the same plants at the rate of 4% (w/w). The bag-treatment method significantly reduced seed damage compared with the untreated control after 6 months of storage (4- to 6-fold decrease in percentage seed damage). Even though relatively low (? 20%), damage by the two insect species controlled by this method was significantly higher than the damage recorded in the seed-treatment method. Orthogonal contrast analysis across test insects showed a significant seed damage difference between treatments with C. ambrosioides and those with L. camara, the latter having the lowest damage profile. The biosafety implication of the use of plant extract-impregnated bags for the protection of food grains against damage by storage bruchid pests is highlighted.     

Resistance of Vigna unguiculata (cowpea) seeds to Callosobruchus maculatus is restricted to cotyledonary tissues

We have investigated the survival of Callosobruchus maculatus larvae when reared on resistant IT81D 1045 Vigna unguiculata seeds, whose resistance has been associated with variant forms of vicilins. Here, we present data which show that larvae of C. maculatus feeding on embryonic axis of resistant cowpea reach a mass of around 28 times higher than those feeding on cotyledonary tissues. Additionally, incorporation of 5–10% of embryonic axis flour on artificial seeds made of resistant flour restores toxicity of seeds to the bruchid. Vicilins purified from both susceptible and resistant embryonic axis had no deleterious effects either on insect development or on insect survival until a level of 4% of incorporation. In contrast, vicilins from resistant cotyledons show an LD₅₀ (50% lethal dose) and WD₅₀ (half weight dose) of 2%. Total vicilin contents of embryonic axes were around two times lower than in cotyledonary tissues, while proteolytic activities of all four proteinase classes were always higher in the former tissues. By 2D‐PAGE we visualized eight protein spots, which seem to be exclusively found on resistant cotyledons. Copyright © 2006 Society of Chemical Industry     

Efficacy of ozone for Callosobruchus maculatus and Callosobruchus chinensis control in cowpea seeds and its impact on seed quality

The insecticidal efficacy of ozone was evaluated against the adults of Callosobruchus maculatus (F.) and C. chinensis L. on stored cowpea seeds under laboratory conditions. Ozone was assessed at concentrations of 0.25, 0.5, 1.0, 1.5, and 2.0 g/m³. The effect of ozone treatments on the adult mortality after 1, 3, 5 and 7 days of treatment, progeny production after 45 days, cowpea seed weight loss, seed germination and chemical constituents of cowpea seeds were determined. Adult mortality of C. maculatus and C. chinensis was improved with the increase in ozone concentration. Thus, all tested concentrations caused complete adult mortality of C. chinensis after 7 days of treatment, while the concentrations of 1.0, 1.5, and 2.0 g/m³ caused complete adult mortality of C. maculatus after the same periods. Progeny of both species was significantly decreased in all ozone concentrations after 45 days of treatment. Nevertheless, strong suppression in progeny production was achieved at the highest concentration of ozone (2.0 g/m³). Also, the ozone treatment at the highest concentration protected the cowpea seeds from damage caused by C. maculatus and C. chinensis for 45 days. In addition, there was no significant effect of ozone treatments on the cowpea seed germination compared with untreated seeds. Moreover, the chemical analysis of treated cowpea seeds showed a slight decrease in protein, fat, carbohydrate, moisture, total phenolics, total flavonoids and tannins contents, and a slight increase in fiber and ash contents compared with untreated cowpea seeds. Our findings suggest the ozone can be effectively used for the control C. maculatus and C. chinensis and can provide sufficient protection of stored cowpea seeds.     

The effect of heating on the haemagglutinating activity and nutritional properties of bean (Phaseolus vulgaris) seeds

The toxic lectins present in red, white and black kidney beans (Phaseolus vulgaris) are sensitive to heat treatment and the efficiency of that treatment is greatly improved by pre-soaking of the seeds. Heating of pre-soaked seeds at all temperatures above 75°C caused a continuous reduction in both their haemagglutinating activity and toxicity. However, the only safe method of eliminating toxicity was to heat the fully hydrated seeds to 100°C for a minimum of 10 min.     

Evaluating the combination of a parasitoid and a predator for biological control of seed beetles (Chrysomelidae: Bruchinae) in stored beans

The two seed beetle species Acanthoscelides obtectus and Callosobruchus maculatus are among the economically most important pests of stored legume seeds world-wide. Insect natural enemies for biological control were considered in IPM strategies to control bean weevils in store. In this contribution, the control effect by a combination of the parasitoid Anisopteromalus calandrae and the predator Xylocoris flavipes on both seed beetle species was evaluated in a laboratory study on black eyed beans Vigna unguiculata. The combination of parasitoid and predator was most effective against both species of seed beetles. However, in C. maculatus the combination was not significantly more effective compared to A. calandrae alone, suggesting a poor effect on adults and no effect on eggs and larvae by X. flavipes. In the case of A. obtectus, the suppressive effect by the combination of the parasitoid and the predator was high. For C. maculatus, germination of black eyed beans was approximately three times higher when both natural enemies were present. The combination of A. calandrae and X. flavipes is considered a promising component for integrated control of A. obtectus.     

A Chitinase and a β-1,3-Glucanase Isolated from the Seeds of Cowpea (Vigna unguiculataL Walp) Inhibit the Growth of Fungi and Insect Pests of the Seed

A chitinase (22000 Da) and a β-1,3-glucanase (26000 Da) were isolated from cowpea ( Vigna unguiculata ) seeds and shown to deter development, in an in vitro assay, of the phytopathogenic fungi Colletotrichum lindemuthianum (agent of anthracnose, a disease of many plants) and Colletotrichum musae (agent of banana peel anthracnose). The isolated chitinase was also shown to negatively affect the development of the cowpea weevil ( Callosobruchus maculatus ) in an artificial seed system. These results suggest that the cowpea seed contains defence proteins that could be, if their levels are properly managed, utilised to promote increased protection of the plant towards attacking fungi and insects.     

Biochemical characterisation of α-amylase inhibitors from Achyranthes aspera and their interactions with digestive amylases of coleopteran and lepidopteran insects

BACKGROUND: Starchy seeds are an important food and a source of dietary ingredients in many countries. However, they suffer from extensive predation by bruchids (weevils) and other pests. α‐Amylase inhibitors are attractive candidates for the control of seed weevils, as these insects are highly dependent on starch as an energy source. RESULTS: A proteinaceous α‐amylase inhibitor from the seeds of Achyranthes aspera was identified, purified and characterised. In electrophoretic analysis, two prominent amylase inhibitor activity bands (AI1 and AI2) were detected. The inhibitor was purified 9.99‐fold with 1206.95 total amylase inhibitor units mg^?1 protein. The molecular weight of the purified inhibitor was around 6 kDa. The isolated α‐amylase inhibitor was found to be resistant to heat and proteolysis. Feeding analysis of Callosobruchus maculatus larvae on a diet containing seed powder of A. aspera revealed that survival of the larvae was severely affected, with the highest mortality rate occurring on the fifth day of feeding. The isolated inhibitor inhibited the majority of amylase isoforms of C. maculatus, Tribolium confusum and Helicoverpa armigera in electrophoretic analysis and solution assays. CONCLUSION: The information obtained in the present investigation could be useful for a genetic engineering approach that would make seeds resistant to storage pest infestations. Copyright © 2011 Society of Chemical Industry     

Effects of cowpea varietal susceptibility and low pressure on the mortality of life stages of Callosobruchus maculatus (Coleoptera: Bruchidae)

Low pressure creates a low-oxygen controlled atmosphere that can kill all developmental stages of Callosobruchus maculatus (F.) (Coleoptera: Bruchidae). This study investigated the interaction of low pressure with seed variety to determine the susceptibility of different cowpea varieties to developmental stages of C. maculatus. Laboratory studies were performed to determine exposure periods to low pressure (32.5 ± 1.0 mm Hg) necessary to kill all life stages of C. maculatus on different cowpea varieties. The results indicated that with the exception of eggs, differences in exposure periods required to achieve 100% mortality varied with life stage and variety. The larval stages were the most susceptible, and larvae required shorter exposure periods to low pressure when reared on C. maculatus resistant varieties (24-125B-3 and IT89KD-288) than on susceptible varieties (CB-3, CB-5, CB-46 and UC-27). Analysis of variance (ANOVA) showed a significant interaction between the life stages of C. maculatus, exposure time and cowpea varieties on mortality. Integration of low pressure applications with storage of cowpea varieties resistant to C. maculatus has the potential of providing an alternative pest management tool to fumigants, which are sometimes used in protecting cowpea in storage from infestation by C. maculatus.     

Characterization of Callosobruchus chinensis (L.) resistance in Vigna umbellata (Thunb.) Ohwi & Ohashi

Resistance to azuki bean weevil, Callosobruchus chinensis, was studied in a series of field and laboratory experiments in two accessions of rice bean (Vigna umbellata (Thunb.) Ohwi & Ohashi), one accession of black gram (V. mungo (L.) Hepper), and one accession of mungbean, (V. radiata (L.) Wilczek). Weevil damage to immature pods of the rice bean accessions, ‘Menaga’ and ‘Miyazaki’, was significantly less than to the susceptible mungbean, VC1973A. In mature pods, the pest damage to the pod wall of Menaga was significantly higher than to VC1973A, whereas the damage to Miyazaki was similar to VC1973A. Seeds within the pods of both rice bean accessions were resistant no matter when the pods were harvested. When the insects were exposed directly on dry seeds, both rice bean accessions and a black gram accession VM2164 were resistant to them. In artificial seeds made by mixing flour of the individual resistant Vigna accessions with VC1973A and subsequently exposed to bruchid oviposition, the higher the quantity of resistant Vigna flour the lower the number of bruchids that emerged from such seeds. No bruchids emerged from artificial seeds containing crude starch fraction from the three resistant Vigna accessions when such seeds were exposed to bruchid infestation, whereas many insects emerged from the seeds containing starch of VC1973A or flour of VC1973A alone. In artificial seeds made by mixing crude protein fractions of the three resistant Vigna accessions with flour of VC1973A, as the concentration of protein increased the number of C. chinensis adults that emerged decreased. Fractionation of crude proteins into acetone-precipitable proteins and peptide and amino acid portions resulted in the loss of antibiosis effect. Artificial seeds made from purified starch-polysaccharides fraction, however, exhibited antibiosis effects if prepared from the rice bean seed of Menaga and Miyazaki but not if made from the black gram seed, VM2164.     

Seed coat phytochemistry of both resistant and susceptible seeds affords some protection against the granivorous beetle Callosobruchus maculatus

The seed coat lies at the interface between the internal structures of the seed and the external environment and thus represents a key arena in the study of seed-herbivore interactions. Callosobruchus maculatus is a cosmopolitan pest of legume seeds, and under post-harvest conditions, females interact directly with the seed testa prior to laying their eggs. Here we investigate the effect of chemical extracts of the seed coat of the resistant Phaseolus vulgaris and the susceptible Vigna unguiculata beans on egg laying preferences and larval development of C. maculatus. Seed coat extracts contained phenolic, glycoside and alkaloid compounds. Upon re-incorporation of extracts into artificial host beans it was found that that several seed coat extracts from both the resistant and susceptible varieties reduced female oviposition and disrupted larval growth and development. However, none of the extracts assayed resulted in complete ovipositional or developmental failure suggesting that complete resistance in P. vulgaris is derived from other physical or chemical properties of the seed and/or seed coat that function either alone or synergistically. Further work is required to elucidate the importance of synergistic interactions between different physiological defence mechanisms on overall plant (seed) resistance.