Effect of field infestations of immature pigeonpea (Cajanus cajan (L.) Millsp.) pods on production of active (flight) and sedentary (flightless) morphs of Callosobruchus chinensis (L.)

A study conducted in Uganda on Callosobruchus chinensis showed that adults emerging from dry seed-infested and immature pod-infested pigeonpea differed in a number of morphological and biological characteristics in one or both sexes. Adults emerging from dry seed-infested pigeonpea were more or less uniform in a number of these characteristics; in contrast, adults emerging from green pod-infested pigeonpea differed depending upon when they emerged: the differences were greatest between those emerging during the 2nd to the 6th days from first emergence, and those emerging from the 8th to the last days of emergence. In one or both sexes, these differences were expressed in terms of elytral hue and size, body length, distance between the compound eyes, length of pygidium, body water content, pre-maturation periods, fecundity and adult longevity. We suggest that these differences are due to polymorphism, and specifically the predominance of flight forms among late emergers from pod-infested pigeonpea. The differences observed in the two morphs of C. chinensis and ascribed to the flight and flightless forms, are thought to be adaptations for survival and propagation under field and storage conditions, respectively. The predominance of emerged adults of the flightless forms from dry seed and the flight forms from green pods were associated with low moisture content in dry seed and high moisture content in green pods, respectively. These observations show that bruchid control measures are needed in the field as well as in storage.     

Resistance to Callosobruchus maculatus (F.) (Coleoptera: Bruchidae) in pigeonpea (Cajanus cajan (L.) Millsp.) and other Cajanus species

Seeds of 24 accessions of pigeonpea (Cajanus cajan (L.) Millsp.) and four other species of Cajanus (formerly Atylosia) were evaluated for their resistance to infestation by Callosobruchus maculatus (F.). None of the pigeonpea accessions was resistant but resistance was evident in three species of Cajanus. In C. platycarpus most of the larvae failed to enter the hard seed coats but the few which did enter the seeds, developed normally. Adults did not emerge from the seeds of C. scarabaeoides, even though most of the larvae entered the seeds. In A. sericeus, the number of larvae entering the seeds as well as adult emergence was significantly reduced.     

Technique to isolate Callosobruchus chinensis (L.) (Coleoptera: Bruchidae) eggs

A simple technique to isolate the eggs of Callosobruchus chinensis from the surface on which they are laid is described. An aqueous extract of their natural food, green gram, Vigna radiata, is used to coat glass beads that attract adult females for oviposition. The thin layer of extract on the glass beads allows separation of the eggs from the bead surface after a 15–20 s immersion in water. When transplanted onto fresh grain treated with glue, the separated eggs exhibit normal development and up to 97% hatching.     

Intraspecific and interspecific competition in Callosobruchus maculatus (F.) and Callosobruchus subinnotatus (Pic) on stored bambara groundnut, Vigna subterranea (L.) Verdcourt

Intraspecific competition was studied in Callosobruchus maculatus and Callosobruchus subinnotatus. Interspecific competition between the two bruchids was also studied to determine which of these species is likely to cause more damage to stored bambara groundnuts, Vigna subterranea in cases of joint infestation. Results showed that increasing the adult density up to 8 females per 10 g of bambara groundnut seeds did not significantly reduce the mean number of eggs laid per female, the number of eggs developing to the adult stage, or the weight of emerged adults of either species. The developmental period of the two species was also not significantly affected. The adult emergence curve of C. maculatus was similar to that of C. subinnotatus and was of the scramble type. C. maculatus performed better than C. subinnotatus in interspecific competition and it achieved this through a higher egg-laying ability and a higher rate of progeny production coupled with a shorter life-cycle. The implications of these findings with respect to damage and possible loss of stored bambara groundnut are discussed.     

Effect of constant temperature and humidity on oviposition and development of Callosobruchus maculatus (F.) and Callosobruchus subinnotatus (Pic) on bambara groundnut, Vigna subterranea (L.) Verdcourt

The effect of different conditions of temperature and humidity on oviposition and development of Callosobruchus maculatus and Callosobruchus subinnotatus was investigated in the present study. Four temperatures (25°C, 30°C, 35°C and 40°C) and three humidities (30%, 60% and 90% r.h.) were evaluated on both species in pure and mixed populations on bambara groundnut (Vigna subterranea) seeds. In both pure and mixed populations, temperature influenced oviposition and progeny development of both species significantly more than humidity. Egg-laying and progeny development in C. maculatus were optimal at 35°C. In contrast, oviposition and progeny development in C. subinnotatus were optimal at 30°C. In mixed populations, where eggs were laid by females of C. maculatus and C. subinnotatus held together, more C. maculatus adults than C. subinnotatus adults were produced at the various temperatures and humidities. No adult progeny of either species developed in pure or mixed populations at 40°C. C. maculatus was more fecund than C. subinnotatus and development from egg to adult took much longer in C. subinnotatus than in C. maculatus in both pure and mixed populations. The implications of these findings for the development and survival of both species in the different agroecologies and seasons in Nigeria are discussed.     

Simulation studies on the effects of solar heat on egg-laying, development and survival of Callosobruchus maculatus (F.) and Callosobruchus subinnotatus (Pic) in stored bambara groundnut Vigna subterranea (L.) Verdcourt

The effects of simulated solar heat on oviposition, development and survival of Callosobruchus maculatus (F.) and Callosobruchus subinnotatus (Pic) in stored bambara groundnut, Vigna subterranea (L.) Verdcourt, were evaluated at three high temperatures (40°C, 45°C and 50°C) at a constant, low humidity (30% relative humidity). Exposure to these temperatures for 6 h significantly reduced oviposition in C. maculatus and C. subinnotatus females. Females of both species that were exposed to 50°C laid significantly fewer eggs than those exposed to 40°C; and in the case of C. maculatus, females exposed to 45°C also laid significantly fewer eggs than those exposed to 40°C. The percentage of eggs laid by females of both species that reached adulthood after exposure to 50°C for 2–6 h was significantly lower than the percentage that developed from eggs laid by females that were exposed to 40°C. No adult developed from eggs of C. maculatus exposed for 6 h at 50°C or from eggs of C. subinnotatus exposed for 2 h at this temperature. For both species, no adult progeny subsequently emerged from seeds harbouring first instar larvae when exposed at 50°C for 2, 4 or 6 h. Older larvae of C. maculatus were more tolerant of exposure at 50°C: 26.8, 10.2 and 0.9% of late instar larvae exposed for 2, 4 and 6 h, respectively, developed to the adult stage. In contrast, no adults of C. subinnotatus emerged from seeds harbouring late instar larvae when exposed at 45°C for 6 h nor in seeds exposed to the temperature of 50°C for 4 or 6 h. On average, immature stages of C. subinnotatus were more susceptible to heat treatment than those of C. maculatus.     

Cajanuslactone,a new coumarin with anti-bacterial activity from pigeon pea [Cajanus cajan (L.) Millsp.] leaves

The bioassay-guided fractionation of the CHCl₃ extract from pigeon pea leaves led to the isolation of a new natural coumarin, named as cajanuslactone (1), together with two known phytoalexins, pinostrobin (2) and cajaninstilbene acid (3). The structure of 1 was elucidated by interpreting spectroscopic data of UV, IR, MS, 1D and 2D NMR. It was determined as 7-hydroxy-5-O-methyl-8-(3-methyl-2-butylene)-4-phenyl-9,10-dihydro-benzopyran-2-one. The anti-bacterial activity against three Gram-positive microorganisms was evaluated in vitro. It was found the new compound cajanuslactone possessed good anti-bacterial activity against Staphylococcus aureus (ATCC 6538), and the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) were 0.031 and 0.125 mg/ml, respectively. The result indicated that the leaf part of pigeon pea is an excellent source of natural anti-bacterial materials. Moreover, cajanuslactone is a potential anti-bacterial agent against Gram-positive microorganisms.     

Biological activity of Cassia occidentalis L. against Callosobruchus maculatus (F.) (Coleoptera: Bruchidae)

In developing countries, traditional control methods are commonly used against stored-product insects and mites. In Senegal, the leaves of Cassia occidentalis L. (Caesalpiniaceae) are used to protect cowpea seeds (Vigna unguiculata L. (Walpers)) against Callosobruchus maculatus (Coleoptera: Bruchidae). The biological activity of the leaves, the seeds and oil of C. occidentalis was evaluated in controlled conditions (28 ± 2°C, 45 ± 5% r.h.) against C. maculatus. At the rate of 10% (w/w), both fresh and dry leaves as well as whole and ground seeds had no contact toxicity on the cowpea beetle. In contrast, seed oil induced an increase in mortality of C. maculatus eggs and first larval instar at the concentration of 10 ml/kg cowpea. The basis of the ovicidal and larvicidal activities are discussed in this paper. Several trials using pure compounds have highlighted that several fatty acids (linoleic, oleic and stearic) are responsible for C. occidentalis toxicity. C. occidentalis seed oil did not reduce the oviposition of C. maculatus at 10 ml/kg seed.     

Efficacy of protectants against Callosobruchus phaseoli (Gyll.) and C. maculatus (F.) (Coleoptera: Bruchidae) in mungbeans

The organophosphorus (OP) compounds chlorpyrifos-methyl, fenitrothion, methacrifos and pirimiphos-methyl, synthetic pyrethroids (SP) deltamethrin and permethrin, and the insect growth regulator (IGR) fenoxycarb were evaluated as protectants against Callosobruchus phaseoli (Gyll.) and C. maculatus (F.) in mungbeans (Vigna radiata (L.) Wilczek). Treated mungbeans were sampled for efficacy testing using C. phaseoli after various periods of storage in hot and humid conditions (30°C, 70% r.h.). Minimum effective application rates were determined for different periods of protection. During storage the OP compounds (chlorpyrifos-methyl, fenitrothion, methacrifos and pirimiphos-methyl) lost activity against C. phaseoli much faster than the SPs (deltamethrin and permethrin) and the IGR (fenoxycarb). Tests were carried out on mungbeans which were infested with C. phaseoli before treatment with pirimiphos-methyl, fenoxycarb or deltamethrin. These tests showed that immature insects in the treated beans were protected from insecticides. Finally, freshly-treated mungbeans were exposed to C. maculatus for comparison with the results for C. phaseoli. The four OP compounds, two SPs and the IGR tended to have similar potency against C. maculatus and C. phaseoli.     

Biology of an egg parasite of Callosobruchus maculatus (Fab.) (Coleoptera: Bruchidae)

Uscana mukerjii (Mani) (Chalcidoidea, Trichogrammatidae) parasitizes eggs of Callosobruchus maculatus. Mated adult U. mukerjii lived for 5.0 ± 0.8 and 4.7 ± 0.9 days (males and females, respectively) at 27 °C. The female copulated only once, whereas males did so up to 4 times. The maximum parasitization of host eggs took place within the first 24 hours of egg laying and declined thereafter. The parasite completed its development inside the host egg in 7.2 days at 27 °C and 60–63% r.h. and emerged after cutting a circular hole in the chorion of the host egg. Mated and unmated females laid 35.4 ± 1.4 and 36.0 ± 2.2 eggs in their life time of which 27.3 ± 1.8 and 23.2 ± 2.8, hatched respectively. Parasitized host eggs did not hatch and were ultimately destroyed. The degree of parasitization varied with the pulses on which the host eggs were laid. This egg parasite may be useful in controlling the populations of C. maculatus and thus help in reducing the damage caused.     

Ethanol modified supercritical fluid extraction and antioxidant activity of cajaninstilbene acid and pinostrobin from pigeonpea [Cajanus cajan (L.) Millsp.] leaves

Supercritical fluid carbon dioxide (SF-CO₂) extraction (SFE) of cajaninstilbene acid (CSA) and pinostrobin (PI) from pigeonpea leaves and antioxidant activity were investigated for the first time. SFE gave higher extraction yields than conventional heat-refluxing extraction (HRE) because of the severe breakage of cell morphology caused by SF-CO₂, which was observed by scanning electron microscopy (SEM). CO₂ flow rate, modifier composition and liquid to solid ratio were firstly optimised, 12 kg/h, and 80% EtOH with a liquid to solid ratio of 10 ml/g raw material exhibited better extraction performance. Then a central composite design (CCD) combined with response surface methodology was used to study the effects of extraction pressure, temperature and time on the extraction yields of CSA and PI, the optimal parameters were 30 MPa, 60 °C and 2 h. An antioxidant activity investigation showed SFE extracts exhibited better free radical-scavenging activity than HRE extracts with an IC₅₀ value of 0.2259 mg/ml.     

Toxicity of dietary avidin and streptavidin to the cowpea bruchid, Callosobruchus maculatus (F.)

Avidin, a large protein from egg whites, powerfully binds biotin, a vitamin for many insects. When avidin was incorporated into the diets of larval Callosobruchus maculatus (F.), the cowpea bruchid, at relatively low levels (10–30 ppm), there was a marked dose-dependent increase in mortality, as well as a small increase in the developmental time of survivors. Avidin toxicity was prevented when biotin was added to the diet together with avidin. Sub-lethal doses of avidin caused reductions in fecundity. Avidin had no effect on the larval feeding rates during the first three instars, even when the larvae were consuming amounts of the protein that would later cause death. In the fourth instar there was a dose-dependent reduction in the rates of feeding as the avidin level in the food increased. Death of the C. maculatus larvae usually occurred at the pupal/adult stage still within the host seed. Streptavidin, a biotin-binding protein from a bacterial source, had effects similar to chicken egg avidin.     

Proteinaceous Inhibitors of Trypsin and of Amylases in Developing and Germinating Seeds of Pigeon Pea ( Cajanus cajan(L) Millsp)

Inhibitors of trypsin and amylase in the extracts of developing seeds of 12 pigeon pea cultivars were analysed using a gel-X-ray film contact print technique and an enzyme-inhibitor assay, respectively. The inhibitors of amylase and trypsin in the extracts of germinating seeds of a pigeon pea cultivar (BDN2) were also studied. Nine trypsin inhibitor bands were detected in mature seeds of all the 12 cultivars. Inhibitory activities against amylase and trypsin were not detected in the extracts of seeds collected 11 and 27 days after flowering (DAF) by the enzyme-inhibitor assay. However, up to three trypsin inhibitor bands could be detected in the extracts of seeds collected 27 DAF by the gel-X-ray film contact technique. Two new slow-moving trypsin inhibitor bands were detected in the extracts of germinating seeds of BDN2 cultivar. These bands were prominent in extracts of seeds 10 days after germination (DAG). The amylase inhibitors and trypsin inhibitors in pigeon pea seeds are late synthesised proteins, their highest levels were observed in mature seeds and they were found to be slowly degraded during germination. Significant inhibitor activities were observed even 15 DAG. The amylases in developing seeds are insensitive to endogenous inhibitors.     

Efficacy of essential oil of Ocimum basilicum L. and O. gratissimum L. applied as an insecticidal fumigant and powder to control Callosobruchus maculatus (Fab.) [Coleoptera: Bruchidae]

Essential oils from sweet basil, Ocimum basilicum, and African basil, O. gratissimum, (Labiatae) grown in Guinea were obtained by steam distillation. Following exposure of newly emerged adult beetles (Callosobruchus maculatus) to 12 h of fumigation using pure essential oils at a dose of 25 μl/vial, 80% mortality was recorded for O. basilicum, 70% for O. gratissimum and 0% in the control. A significant difference was observed between the responses of males and females with males exhibiting greater sensitivity. When 1 g of aromatized powder was applied to adults, a 50% lethal concentration at 48 h was found to be 65 μl/g for O. basilicum and 116 μl/g of O. gratissimum oils. The essential oils from the two plant species exhibited a significant effect both on the egg hatch rate and on the emergence of adults. The egg hatch rate was reduced to 3% with O. basilicum and 15% with O. gratissimum using an essential oil concentration of 30 μl, whereas the egg hatch rate for the control was 95%. When compared with the control (97%), adult emergence dropped to 0% with O. basilicum and to 4% with O. gratissimum. Storage bioassays were run to assess the long-term effect of powders aromatized with essential oils of Ocimum. Complete protection was observed over 3 months starting at a dose of 400 μl in the case of both oils. From a germination test, it was concluded that aromatized powders have no significant effect on the seed germination rate. After 5 d, a rate of 88% germination was seen in seeds treated with aromatized powder and protected from insects, compared with 97% for untreated seeds that were not exposed to insects.     

Characterization and insecticidal properties of globulins and albumins from Luetzelburgia auriculata (Allemao) Ducke seeds towards Callosobruchus maculatus (F.) (Coleoptera: Bruchidae)

We evaluated the development of Callosobruchus maculatus growing in artificial seeds composed of Vigna unguiculata (cowpea) seed flour mixed with exogenous proteins from Luetzelburgia auriculata. Albumin and globulin fractions from Luetzelburgia auriculata were characterized in terms of protein content, amino acid composition and antimetabolic proteins (trypsin/chymotrypsin inhibitory, porcine pancreatic alpha-amylase inhibitory, lectin activity and presence of chitin-binding proteins). Both fractions were distinct in terms of protein content and diversity as determined by electrophoresis. Lectin activity was present only in the globulins. Neither fraction exhibited inhibitory activity towards porcine pancreatic alpha-amylase but trypsin inhibition was observed. Interestingly, chitin-binding proteins were detected in both protein classes. Albumins had a severe effect upon larval development and were detrimental to insect emergence (LD₅₀=0.4%) while globulins displayed slight toxicity upon larval development and no effect towards insect emergence. The presence of serine proteinase inhibitory activity and chitin binding proteins could explain, at least in part, the harmful effects on C. maculatus development while lectin activity and amino acid availability seem not to correlate with any deleterious effects. Luetzelburgia auriculata would be an interesting source of seed proteins to study behavior of C. maculatus upon infestation and genes coding for insecticidal proteins could become candidates for molecular biology programs devoted to producing transgenic seeds expressing resistance towards the beetle.     

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.     

Enzyme assisted extraction of luteolin and apigenin from pigeonpea [Cajanuscajan (L.) Millsp.] leaves

Luteolin and apigenin are naturally occurring flavones with a wide spectrum of pharmacological properties. In the present study, enzyme assisted extraction of luteolin and apigenin from pigeonpea leaves using commercial plant cell wall degrading enzyme preparations including cellulase, beta-glucosidase and pectinase were examined. We found that pectinase offered a better performance in enhancement of the extraction yields of luteolin and apigenin than cellulase and beta-glucosidase. The pectinase assisted extraction process was further optimized by varying different parameters such as pectinase concentration, time of incubation, pH of pectinase solution, and incubation temperature. The optimum parameters were obtained as follows: 0.4 mg/ml pectinase, incubation for 18 h at 30–35 °C, pH of pectinase solution 3.5–4. Under the optimum conditions, the extraction yields of luteolin and apigenin achieved 0.268 and 0.132 mg/g in pectinase treated sample, which increased 248% and 239%, respectively, compared with the untreated ones.     

Varietal effects of cowpea, Vigna unguiculata, on tolerance to malathion in Callosobruchus maculatus (Coleoptera: Bruchidae)

The impact of cowpea variety on the response of cowpea bruchid, Callosobruchus maculatus, to malathion was investigated. The interaction of six cowpea varieties (Adamawa Brown, Ife BPC, Ife Brown, Lilongwe, Ntcheu and NCRI-L25) with the geographical strains of C. maculatus (Brazil and Cameroon), temperature (23, 25, 27 °C) and insecticide concentration were considered. Cowpea variety (V) had an unpredictable effect on C. maculatus response to malathion. Bruchid populations produced by Ife BPC were the most susceptible to malathion while those yielded by NCRI-L25 were the most tolerant. Regardless of the cowpea variety, the Brazil strain showed higher tolerance than the Cameroon strain. There was significant effect of temperature (T) and insecticide concentration (C) on malathion tolerance in both strains (S). Likewise, there was significant impact of all two-way interactions on cowpea bruchid tolerance except V x C. Significant three-way interactions on C. maculatus tolerance to malathion was only observed in S × T × V and S × T × C. The predictability of changing one of the factors on the susceptibility of C. maculatus to insecticide was very low. This study suggests a need to take the insecticide tolerance of insect populations produced by novel varieties into account during plant breeding in addition to factors such as yield and resistance to insect and disease attack.     

Status of glycemic index of paddy rice grain (Oryza sativa L.) on infestation by storage pest Sitotroga cerealella

The harvested paddy rice grains are normally stored by growers under normal conditions. Paddy rice grains are natural food for storage pests like Sitotroga. cerealella, a major insect that appears during paddy storage. The changes in grain composition due to such insect infestation in relation to human health especially the alteration in their Glycemic Index (GI) value have not been studied so far. The present investigation was aimed at studying the effect of S. cerealella infestation on rice grain quality and glycemic index of stored paddy grains. Five diverse rice genotypes with different quality traits were evaluated for grain quality after S. cerealella infestation. The tolerance to insect and variation of grain quality varied with rice genotype significantly. The glycemic index, glycemic load, total carbohydrate, amylose content and resistant starch were affected to a great extent. The consumption of affected grains may create health issue due to imbalance in nutrition and unhygienic condition. Improper storage of paddy after harvesting and processing results in infestation by pests at a large scale. Thus, the grain quality needs to be evaluated to judge the suitability of affected grain for consumption.