Feeding bioassay for stored-product insect pests using an encapsulated food source

A procedure that was developed to encapsulate liquid and semiliquid diets was used to encapsulate dry diet for use in a feeding bioassay for beetles that are pests of stored products. Vacuum was used to form Parafilm^® into numerous 6 mm diameter wells. The wells were filled with clean sand (control) or ground dry dog food (test), and the Parafilm^® sealed to produce individual pellets. A single pellet was then placed in the center of a 9 cm diameter Petri dish and feeding activity of groups of ten adults of Sitophilus oryzae (L.), the rice weevil; Oryzaephilus surinamensis (L.), the sawtoothed grain beetle; and Tribolium castaneum (Herbst), the red flour beetle, was tested. The number of insects on the encapsulated pellet and the amount of food or sand scattered were checked hourly for the first 8 h of the study and after 24 h. Few insects were observed on pellets containing sand and little or no sand was observed scattered outside of the pellet, so the presence of dog food in the pellet was needed for insects to feed on the pellet. In tests with encapsulated dog food, the amount of food scattered provided a better quantitative measure of feeding than the number of insects on the pellet. Insects that were starved for 48 h caused greater amounts of food scatter than insects starved for 24 h prior to the test. In direct comparisons among all three species, T. castaneum responded the most slowly and the bioassay may be improved by increasing the amount of time starved. Sitophilus oryzae responded very quickly and the entire pellet was essentially consumed within the first 4-5 h of the study. The encapsulated diets provide a promising method to evaluate feeding behavior of stored-product insect pests.     

Susceptibility of stored-product insects to enhanced diatomaceous earth

Mortality of adult Rhyzopertha dominica (F.), Sitophilus oryzae (L.) and Tribolium castaneum (Herbst) was recorded after exposure for different times to application rates of 0, 0.5, 1, 1.5 or 2 g/m² of the diatomaceous earth (DE) DEBBM-P/WP and to 0, 3, 4, 5 or 6 g/m² of the DE Protect-It^®. Mortality of all insects increased with increasing exposure interval; dry dusts were more effective than slurries, and overall mortality was greater for DEBBM-P/WP than Protect-It^®.     

Laboratory assessment of insecticidal effectiveness of natural zeolite and diatomaceous earth formulations against three stored-product beetle pests

The insecticidal effectiveness of two natural zeolite formulations (Minazel plus and Minazel), applied to wheat at selected rates of 0.25, 0.50 and 0.75 g/kg, and a diatomaceous earth formulation (DE) (Protect-It™), applied at the recommended rates of 0.15 g/kg for Sitophilus oryzae, 0.20 g/kg for Rhyzopertha dominica and 0.30 g/kg for Tribolium castaneum, were tested under laboratory conditions (24 ± 1 °C temperature and 45 ± 5% relative humidity). The highest adult mortality was observed after the longest exposure period of 21 days and 7 days of recovery, when all three zeolite dosage rates and the recommended DE dosage caused 97-100% mortality of S. oryzae and 94-100% of T. castaneum. On the other hand, 100% mortality was not achieved in any test variant involving R. dominica; the highest (about 92%) was detected for DE, while 52% and 79% mortality was achieved with the zeolites at the highest rate of 0.75 g/kg. Progeny reduction by >90% was achieved after 21 days of contact of all three beetle pests with DE-treated wheat, while the same level of reduction was achieved for S. oryzae and T. castaenum only after contact with the highest rate of the zeolite product, Minazel. Thus the two zeolite formulations are comparable to diatomaceous earth in controlling adult S. oryzae, R. dominica and T. castaneum, but only the Minazel formulation could effectively protect wheat from attack by S. oryzae or T. castaneum, and only with a higher rate of application than for the DE formulation.     

Detection of eggs of stored-product insects in flour with staining techniques

Chemical stains were investigated with the aim of developing a method for detecting eggs of stored-product insects in refined wheat flour (white flour), so that eggs and flour particles could be distinguished easily. Eggs of the rust-red flour beetle, Tribolium castaneum, the saw-toothed grain beetle, Oryzaephilus surinamensis and the rice moth, Corcyra cephalonica were detected by staining with 0.1% bromocresol green or 1:1 mixture of bromocresol green+0.5% orange G solution after acid digestion of the flour particles. In contrast to the standard AACC method of staining with iodine solution where both eggs and flour particles were stained brownish-yellow or yellow, bromocresol green alone or in mixture with orange G, differentially stained the insect eggs and the flour particles distinct orange and green, respectively. Eggs in flour could also be detected using 0.05% acid fuchsin but both eggs and flour particles were stained uniformly pink.     

Effect of “soft-electron” (low-energy electron) treatment on three stored-product insect pests

Developmental stages of three stored-product insect pests viz. Tribolium castaneum (Herbst), Plodia interpunctella (Hübner) and Callosobruchus chinensis (L.) were exposed to “soft electrons”, low-energy electrons. Soft electrons at an acceleration voltage of 170 kV effectively inactivated eggs, larvae and pupae of T. castaneum and P. interpunctella. The adults of T. castaneum and P. interpunctella were inactivated by treatment for 10 min (4.8 kGy) and 15 min (7.2 kGy), respectively. Soft electrons at 170 kV also inactivated the eggs of C. chinensis effectively. The adults of C. chinensis survived a 15 min exposure (7.2 kGy), but were inactivated having lost the ability to walk after a 5 min exposure (2.4 kGy). Soft electrons at 170 kV could not completely inactivate the larvae of C. chinensis inside beans, because the electrons with low penetration did not reach the larvae due to the shield of beans. The movement of T. castaneum larvae exposed to soft electrons was sluggish for several days, and then the larvae died with their hindguts emerged from their posterior ends 17 days after the treatment. DNA comet assay of cells of P. interpunctella larvae indicated that soft electrons at 170 kV damaged DNA in the larvae.     

A simple technique to assess compounds that are repellent or attractive to stored-product insects

We have developed a simple and rapid technique that mimics storage conditions, and determines if products are repellent or attractive to stored-product insects. The technique determines the response of insects to potential repellents and attractants by measuring their movement from grain. The technique used a device consisting of a perforated cup (2 mm perforations) that holds 200 g of wheat. A Petri dish and cup collected the insects as they left the wheat. Several natural products were tested for repellency: diatomaceous earth (DE), ground peas (Pisum sativum), protein-rich pea flour, pea starch, and pea fibre. Adult insects of three species were tested: the rice weevil, Sitophilus oryzae, the red flour beetle, Tribolium castaneum, and the rusty grain beetle, Cryptolestes ferrugineus. DE at 0.01% was repellent to all insects tested. Pea fibre, pea protein, and ground pea at 1% caused increased emigration of C. ferrugineus from the wheat. Pea starch did not affect movement out of the grain for all three insects. Only pea fibre and ground pea increased the movement of T. castaneum out of the grain. For S. oryzae, there were no differences after 1 h, but after 24 h both pea protein and ground pea increased movement out of the grain. Several potential attractants were placed outside the grain and the emigration out of the grain noted. For R. dominica, the commercial R. dominica pheromone increased the emigration of insects from the grain; R. dominica adults on broken grain enclosed in a ventilated vial in the collection jar also increased emigration, but not as much as the synthetic pheromone. The commercial Tribolium pheromone did increase movement out of the grain for T. castaneum, but the other treatments were no different from the control. None of the potential attractants increased the movement of S. oryzae from the grain. The implications of this work are discussed with reference to controlling and sampling stored-product insect pests.     

Studies on insect infestation in chocolates

The ability of stored-product pests including the cigarette beetle, Lasioderma serricorne, the sawtoothed grain beetle, Oryzaephilus surinamensis, the rust-red flour beetle, Tribolium castaneum, and the almond moth, Ephestia cautella, to infest chocolates under packaged and unpackaged conditions was investigated in the laboratory at 25±1 °C and 65±5% r.h. Four types of chocolates were investigated: milk, nut, dried fruit and nut, and wafer chocolates. Adults (beetles only, 20 per replicate) or eggs (30 per replicate) were released on unpackaged and packaged chocolates and infestation levels (number of living adults and larvae) were determined 45 days later. When adult beetles were released on unpackaged chocolates, the degree of infestation varied depending on the species and the type of chocolate. The highest infestation observed in unpackaged chocolate was that of O. surinamensis in wafer chocolate (mean 138.4). When eggs were released on unpackaged chocolates, the most numerous species was E. cautella in dried fruit and nut chocolate (mean population=180.8). With packaged chocolates exposed to adults or eggs, insect infestation was nil or negligible (mean population <6.0). Although infestation levels were low, infestations were found in 50% of treatments over all. Damage to the packaging material along the folds or edges was observed in infested chocolates. The study has shown that milk, nut, dried fruit and nut, and wafer chocolates can support insect infestation and therefore, insect-proof packing of the chocolates and storage under hygienic conditions are important to avoid customers’ complaints.     

Insect population dynamics in commercial grain elevators

Data were collected in 1998-2002 from wheat stored in commercial grain elevators in south-central Kansas. Bins at these elevators had concrete walls and were typically 6-9 m in diameter and 30-35 m tall. A vacuum-probe sampler was used to collect grain samples in the top 12 m of the wheat in each bin. The primary insect species found in the wheat samples were: Cryptolestes ferrugineus, Rhyzopertha dominica, and Tribolium castaneum. In the top 3.7 m of grain, R. dominica, C. ferrugineus, T. castaneum and Sitophilus oryzae made up 44, 36, 19 and 1% of the insects found in the samples, respectively. From 3.8 to 12.2 m, R. dominica, C. ferrugineus, T. castaneum and S. oryzae were present at 84, 8, 8, and 1%, respectively. The most prevalent species also changed over time. In June, the start of wheat harvesting and storage in Kansas, insect density was low in the bins. At this time, C. ferrugineus was the most common insect, and it was found mostly in the top grain sample (0-1.2 m). In September through November, C. ferrugineus and R. dominica were at similar densities; however, from February to March, R. dominica was more common.Generally, insect density was greatest at the top and decreased with grain depth. Very few insects were found in samples collected from greater than 12 m (most of the bins contained grain to depths of 24-36 m). Insect density for all species increased rapidly from June through October. During this period less than 20% of the bins had economically significant insect densities (>2 insects/kg). From October until February, the average insect density remained fairly constant but it was greatly reduced in April, May, and June. Bins that had insect densities >2 insects/kg tended to be located adjacent to other heavily infested bins.     

Effect of low pressures on the survival of cocoa pests at 18°C

This study forms part of an effort to eliminate the need for fumigation with methyl bromide to control insect infestations in stored cocoa beans, through development of novel alternative vacuum-hermetic technology. In this communication, the effects of low pressures and exposure time were studied on the mortality of insects at a temperature of 18°C, chosen to simulate cocoa bean storage conditions in temperate climates.Three insect species were used, two of which are major pests of cocoa beans in producer countries, Ephestia cautella (Walker), and Tribolium castaneum (Herbst), while the third, Oryzaephilus surinamensis (L.), is a potential storage pest in temperate climates. For T. castaneum and E. cautella the egg stage was the most resistant to 55±10 mm Hg at 18°C, the times needed to obtain 99% egg mortality were 96 and 149 h, respectively. For O. surinamensis, the adult stage was the most resistant with 164 h being required to obtain 99% mortality.     

Interspecific associations among stored-grain beetles

Recent increases in prices of raw grain, including wheat, will reduce action thresholds for insect damage and therefore justify more research into management practices and understanding of pest ecology in stored grain. Compared to most other habitats, natural or man-made, a filled grain silo constitutes a unique and fairly homogeneous habitat in which food availability for many grain-feeding insects is unlimited. A fundamental aspect of stored-grain insect ecology is a better understanding of associations among common beetle species. We analyzed the densities of three important stored-grain beetle species, Rhyzopertha dominica (F.) (Coleoptera: Bostrichidae), Cryptolestes ferrugineus (Stephens) (Coleoptera: Laemophloeidae), and Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) in wheat samples collected in 1999–2001 from 129 grain silos in Kansas. The beetles studied here are highly mobile, and the number of insects in each grain sample is a result of the beetles' preference for favorable micro-environmental conditions and possibly of intra- and interspecific associations. In general, the number of T. castaneum in a grain sample increased as the number of R. dominica increased, but the number of C. ferrugineus was not correlated with the number of R. dominica. The densities of both T. castaneum and R. dominica decreased as the number of C. ferrugineus increased. Cryptolestes ferrugineus and T. castaneum can be predators and the species composition of insects in a grain sample may be modified by predation. As T. castaneum populations increased, so did R. dominica but not C. ferrugineus. Our analysis of the species composition in grain samples is discussed in an ecological context.     

Survival and reproduction of Tribolium castaneum (Herbst), Rhyzopertha dominica (F.) and Sitophilus oryzae (L.) following periods of starvation

This study determined the starvation tolerance of Tribolium castaneum (Herbst), Rhyzopertha dominica (F.) and Sitophilus oryzae (L.) in terms of both adult survival and reproduction, the impact of starvation on reproduction not having been studied before. Experiments were conducted at 30 °C and 55% or 70% r.h. using a laboratory strain and a field strain of each species. The number of progeny was a better indicator of the impact of starvation on a species than adult survival. Tribolium castaneum was the most tolerant species, requiring up to 35 d starvation before no progeny were produced. Rhyzopertha dominica and S. oryzae required up to 8 d starvation before no progeny were produced. The results suggest that hygiene will have a greater impact on populations of S. oryzae and R. dominica than T. castaneum.     

Spatial distribution of some beetles infesting a feed mill with spatio-temporal dynamics of Oryzaephilus surinamensis, Tribolium castaneum and Tribolium confusum

Food-bait traps were used to study spatial and temporal distribution of the coleopteran fauna of the first floor of a feed mill in Central Italy. A total of 3396 beetles were captured during the 1-year survey. Beetles were most abundant during July and August and least abundant during February and March. Tribolium confusum du Val was the most abundant and widespread, followed by Oryzaephilus surinamensis (L.), Tribolium castaneum (Herbst), Attagenus brunneus Faldermann, Sitophilus oryzae (L.), and Stegobium paniceum (L.). The spatial patterns of annual catches of these six species were depicted by contour maps. In addition, high catches of O. surinamensis, T. castaneum and T. confusum, allowed construction of contour maps for monthly trap catches. The populations of A. brunneus were located in the bagging, milling, formulation and pelleting sites, and in the area around the conveyer belt. The highest populations of O. surinamensis were in the entry zone of the unloading pit and in the area around the conveyer belt with finished products. Sitophilus oryzae were present near the entry door of the unloading pit room, around the storage bins and in the storeroom. Populations of S. paniceum were found in the unloading pit as far as the conveyer belt and in a corner of storeroom. Tribolium castaneum was limited to a localized area in the room where raw material was processed and in the entry to the unloading pit room. The highest T. confusum populations were located in the milling, formulation and pelleting sites, and near balance and storage bins. Comparison of the spatio-temporal dynamics of these pests showed a segregation of populations, in both time and space, and a strong interaction among species is suggested. On the basis of our results, the spatial and temporal distributions are significantly affected by various factors, such as food availability, processing practices and temperature conditions in different areas.     

Feeding deterrent activity of terpenoid lactones with a p-menthane system against stored-product pests

Feeding deterrent activity of eight enantiomeric pairs and one optically inactive terpenoid lactone with a p-menthane system against three storage pests was determined. The lactones were tested on adults of Sitophilus granarius, adults and larvae of Tribolium confusum and larvae of Trogoderma granarium. The isomeric starting natural compounds, (+) and (−) pulegones and (+) and (−) isopulegols, were also tested. The results showed that the introduction of the lactone moiety into the p-menthane system produced antifeedant activity in the lactones obtained. The lactones with a spiro arrangement of lactone and cyclohexane rings were more active than those with condensed rings. The configuration of chiral centres present in the molecule significantly influenced the activity of the compounds studied. In most cases, lactones obtained from R-(+)-pulegone were more active antifeedants than those obtained from S-(−)-pulegone.     

Toxicity of ethyl formate to adult Sitophilus oryzae (L.), Tribolium castaneum (herbst) and Rhyzopertha dominica (F.)

Fumigations were conducted using a continuous flow-through laboratory process to maintain constant concentrations of ethyl formate and low levels (<0.8%) of respiratory carbon dioxide. The procedure minimised the effects of sorption by exposing test insects without media and minimised the effect of carbon dioxide by use of continuous flow. The concentration×time (Ct) products of ethyl formate for adult Sitophilus oryzae, Tribolium castaneum and Rhyzopertha dominica at 25 °C and 70% relative humidity for the 6 h exposure were, respectively: (1) LD₅₀ 107.8, 108.8 and 72.8 mg h L⁻¹ and (2) LD_99.5 207.4, 167.1 and 122.2 mg h L⁻¹. Endpoint mortality was reached within 24 h of initial exposure.     

Integrated management of insect vectors of Aspergillus flavus in stored maize, using synthetic antioxidants and natural phytochemicals

The purpose of this study was to investigate the insecticidal activity of two benzoic acids 2(3)-tert-butyl-4 hydroxyanisole (BHA) and 2,6-di(tert-butyl)-p-cresol (BHT); two phenolic acids 3-phenyl-2-propenoic acid (CA) and trans-4-hydroxy-3-methoxycinnamic acid (FA) and two essential oils of Eugenia caryophyllata (clove tree) and Thymus vulgaris (thyme) against Sitophilus zeamais, Tribolium confusum and Rhyzopertha dominica, vector carriers of aflatoxigenic fungi in stored maize. The susceptibility of insects, the frequency of isolation of Aspergillus section Flavi in insects and maize, and the analysis of aflatoxin B₁ in maize were determined. BHA, BHT, BHA/BHT mixture and the natural phytochemicals AF and AF/AC mixture showed the highest insecticidal activity against S. zeamais, T. confusum and R. dominica after 120 days of incubation. The insecticidal efficacy of the volatile fraction of essential oils of clove and thyme showed less inhibition. There was no contamination of Aspergillus section Flavi in dead and live insects collected from maize treated with BHA. No aflatoxin B₁ accumulation was detected in the control and treatments. The information obtained shows that these substances have the potential to control pest insect vectors of aflatoxigenic fungi in stored maize in microcosms during 120 days.     

Response of mixed-age cultures of phosphine-resistant and susceptible strains of lesser grain borer, Rhyzopertha dominica, to phosphine at a range of concentrations and exposure periods

Mixed-age cultures, containing all life stages, of a highly resistant strain (Strong-R) of lesser grain borer, Rhyzopertha dominica (F.), were exposed to a series of fixed concentrations of phosphine at a range of exposure periods at 25 °C. A susceptible strain and a less-resistant strain (Weak-R) were also tested. The aim was to characterise the resistant strain and determine if it could be controlled with phosphine. Times to population extinction (TPE) were recorded and lethal time (LT_99.9) values calculated. The relationship between exposure period t and phosphine concentration C for the resistant strains were: for Strong-R LT_99.9C^0.5457t=3.852 and TPE C^0.6105t=4.0404 and for Weak-R LT_99.9C^0.3553t=3.6521 and TPE C^0.4507t=3.4833. The results were used to define a range of minimum exposure period ×concentration protocols for control of the Strong-R populations. For example, at 1.0, 0.3, and 0.2 mg l⁻¹ complete control can be expected in 5, 10 and 14 days, respectively. This information will be used to recommend phosphine rates for field trial and eventual registration.     

Impact of differing population levels of Rhyzopertha dominica (F.) on milling and physicochemical properties of sorghum kernel and flour

The effects of different population levels of Rhyzopertha dominica (F.), the lesser grain borer, on physiochemical properties of sorghum kernels and flour, were investigated through a laboratory study at 27 and 32 °C, and 57% relative humidity. Initial population level and temperature, and their interaction, were significant for the number of F₁ progeny and feeding damage (P<0.01). A strong positive correlation was also found between initial population size, number of F₁ progeny, percentage of insect-damaged kernels (IDKs) and feeding damage. The impact of R. dominica on the milling quality of sorghum was seen through a reduction in abrasive hardness, milling yield, and kafirin content. Initial population and temperature affected most pasting properties, and overall pasting viscosity increased with initial population, number of F₁ progeny, and percentage of IDK at 32 °C. Results show that R. dominica can potentially impact the milling quality of sorghum and also alter the physicochemical properties of sorghum flour.     

Response and genetic analysis of malathion-specific resistant Tribolium castaneum (Herbst) in relation to population density

Extensive use of malathion for pest control on stored cereals has resulted in worldwide resistance in red flour beetles, Tribolium castaneum. In this paper we investigate population density effects on the malathion-specific resistance in PRm, a strain from the Philippines, in an integrated resistance management framework. Two populations of malathion-specific resistant (PRm) and a malathion-susceptible strain of T. castaneum were bred at different densities: low (4 adults/g) and high (12 adults/g) density on wheat plus brewer's yeast in the laboratory. After eight generations, slopes of probit regression lines and LC₅₀ values were used to monitor the effect of insect rearing density on the progression of malathion-specific resistance. The LC₅₀ of the malathion-susceptible strain (Asm) did not change significantly during selection while LC₅₀s varied for both the high-density and low-density lines of PRm, the LC₅₀ of malathion ranged from 27.51 to 34.06 and from 21.14 to 29.39 μg malathion cm⁻² for high and low density, respectively.More than 33 generations were required to achieve a 10-fold increase of resistance for the low-density line compared to only 17 generations for the high-density line. Calculations from published formulae suggested that the malathion-specific resistance of both high- and low-density lines was under monofactorial control, with complete dominance.The data showed that environmental factors such as population density differences in insect rearing and development may influence the heritability of resistance. Furthermore, the variability in results published worldwide on resistance emphasises the need to standardize test conditions across laboratories.     

Heat treatment for disinfestation of empty grain storage bins

An alternative to fumigants and insecticides for controlling stored-product insects in empty grain storage bins prior to filling is heat treatment, in which the temperature is quickly raised to a minimum of 50 °C and held there for 2-4 h. Effectiveness of heat treatment on empty grain storage bins was evaluated for five commercial propane and electric heat-treatment systems by measuring air temperature and associated mortality of Tribolium castaneum (Herbst), the red flour beetle, Sitophilus oryzae (L.), the rice weevil, and Rhyzopertha dominica (F.), the lesser grain borer, exposed for different time intervals. Eleven locations, six above and five below the drying floor, were monitored for air temperature and associated mortality of the three insect species, using arenas initially stocked with live adult insects. Data were analyzed separately for each heating system, with floor location and time interval as main effects for insect mortality. A high-output propane heater (29 kW) produced 100% mortality in 2 h for the three insect species at all test locations. An electric duct-heater system (18 kW) also produced 100% mortality at all test locations after 40 h when aided by a complicated interior heat-distribution system. The other three systems produced less than 100% mortality.     

Protection of stored maize from insect pests using a two-component biological control method consisting of a hymenopteran parasitoid, Theocolax elegans, and transgenic avidin maize powder

The hymenopteran parasitoid, Theocolax elegans (Westwood), and transgenic avidin maize powder were tested to determine if their individual or combined use would protect stored grain from infestation by both internal and external insect pests. Small-scale tests were conducted in plastic jars containing 3 kg of non-transgenic maize. We tested treatments of 0.3% powdered avidin maize, the parasitoid wasp, and the combination of the parasitoid plus 0.3% powdered avidin maize. One pair each of Sitophilus zeamais Motschulsky, Tribolium castaneum (Herbst), and Cryptolestes ferrugineus (Stephens) was added to each jar. After 8 weeks, the entire contents of each jar were examined for adult insects. Control and avidin maize powders had no detrimental effects on the beneficial insect parasitoid T. elegans. The parasitoid suppressed populations of the internal feeder S. zeamais. The avidin maize powder treatment had no effect on S. zeamais because these larvae developed inside the maize kernels where no avidin maize powder was present. For S. zeamais, the combination treatment was not significantly different from the parasitoid treatment. In contrast, populations of the external feeder T. castaneum were not suppressed by the parasitoid but were suppressed by the avidin maize powder treatment. The parasitoid-avidin combination treatment produced the greatest percentage reduction for all three insect species and resulted in 78%, 94%, and 70% reductions in populations of S. zeamais, T. castaneum, and C. ferrugineus, respectively, when compared to the control treatment. The percentage reductions for the parasitoid treatment were 70%, 8%, and 20% for S. zeamais, T. castaneum, and C. ferrugineus, respectively. For the avidin maize powder treatment, populations of S. zeamais, T. castaneum, and C. ferrugineus were reduced by 10%, 85%, and 40%, respectively. The combination treatment of avidin maize powder plus the release of parasitoid wasps was superior to either treatment alone when applied to mixed populations of internal and external feeders.