Mechanisms of premating isolation between Helicoverpa armigera (Hübner) and Helicoverpa assulta (Guenée) (Lepidoptera: Noctuidae)

Helicoverpa armigera and Helicoverpa assulta are sympatric sibling species, and in the laboratory they can interbreed and produce viable offspring. To assess the contributions of temporal barriers and sexual barriers to premating isolation, we investigated both the temporal rhythms of calling behavior and pheromone titers of H. armigera and H. assulta females and the behavioral responses of males to conspecific and heterospecific calling females in a wind tunnel. Both H. armigera and H. assulta females called throughout the scotophase, and there was more calling during the second half of the scotophase than during the first half. Maximal pheromone titer and maximal calling activity in H. armigera synchronously occurred at the sixth hour into the scotophase, whereas, in H. assulta, the maximal pheromone titer occurred 2 h before the peak of calling. Pheromone blend ratios of the two species were opposite and, within each species, changes in the ratio within the scotophase and at different ages were relatively small. Males of both H. armigera and H. assulta responded strongly to their conspecific calling females in the wind tunnel and completed the whole courtship sequence. In contrast, they did not land and had no copulation attempts in response to heterospecific calling females. These results show that the two species do not have obvious temporal differences in calling behavior and pheromone production, and the specificity of sex pheromone blend emitted by females plays a key role in their premating isolation. In addition, we summarized the potential isolation mechanisms of H. armigera and H. assulta.     

Modeling Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) damages on cotton

We studied and modeled damage caused by Helicoverpa armigera larvae on cotton with the aim of developing a coupled crop pest model. Two damage components were studied: the voracity (quantity of fresh matter and number of organs consumed) and feeding preferences (type of organ infested). The laboratory no-choice study of voracity on excised squares and bolls revealed that an H. armigera larva consumes 2,856 mg of fresh matter throughout its larval life, with the sixth instar consuming 86% of this quantity. This consumption rate corresponded to 23.6 squares, or 7.8 bolls. We developed equations to predict the quantity of fresh matter uptake from an individual plant organ, according to the organ mass and the larval instar. The field study of feeding preference confirmed previous findings that larvae prefer squares to bolls, with this preference decreasing as the larval age increases. However, no significant relationship was noted between the age of larvae and the size of infested organs within each organ class (square or boll). We developed a logistic model to predict the probability of a larva infesting a boll rather than a square. According to this model, the relative organ availability in the field and the larval instar were found to be significant factors.     

Structure and ultrastructure of the silk glands and spinneret of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

This study provides comprehensive documentation of silk production in the pest moth Helicoverpa armigera from gland secretion to extrusion of silk thread. The structure of the silk glands, accessory structures and extrusion apparatus are reported. The general schema of the paired silk glands follows that found for Lepidoptera. Morphology of the duct, silk press, muscle attachments and spigot are presented as a three-dimensional reconstruction and the cuticular crescent-shaped profile of the silk press is demonstrated in both open and closed forms with attendant muscle blocks, allowing advances in our knowledge of how the silk press functions to regulate the extrusion of silk. Growth of the spigot across instars is documented showing a distinctive developmental pattern for this extrusion device. Its shape and structure are related to use and load-bearing activity.     

Karyomorphological effects of two new oil formulations on Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

The cotton bollworm Helicoverpa armigera is a serious pest of many economically important crops. Since this pest has become resistant to the conventional synthetic insecticides, newer compounds and formulations are being developed against this insect pest. Many natural compounds isolated from the plants were tested against this pest. Among them Hyptis suaveolens and Melochia corchorifolia showed insecticidal properties against H. arnigera. Based on bioefficacy studies, caryophyllene and β-sitosterol were isolated from H. suaveolens and M. corchorifolia respectively. The isolated natural compounds were further developed as formulations in various combinations with neem (Azadirachta indica) and karanj (Pongamia pinnata) oils. The present study indicated that the formulations influenced the karyomorphology of H. armigera.     

Antifeedant activity of numb and salty taste compounds against the larvae of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

Helicoverpa armigera, an important polyphagous insect pest in agriculture, attacks more than 200 plant species of more than 30 families. Our previous study showed that the choice feeding percentages of H. armigera larvae to tobacco leaf discs treated with sweet, bitter, and hot taste substances were higher than the control leaf discs, while numb and salty substances could significantly inhibit their feeding. To quantitatively determine the synergistic effect of numb and salty substances, in this paper, the antifeeding activities of numb and salty substances and their mixtures blended in different doses or volume ratios were assayed on H. armigera larvae. The first bioassay was designed to elucidate the relative feeding preference of the larvae to the leaves from several common host species, each paired with tobacco leaf discs, and the result indicated that the most preferred host leaf by the larvae was tobacco leaf, followed by cotton and peanut leaves, suggesting that tobacco leaf was the most suitable matrix for the antifeeding bioassay, and the larval consumption of maize, pepper, or tomato leaves were significantly lower than that of tobacco leaf. The second bioassay was to test the choice feeding response of H. armigera larvae to tobacco leaf discs treated with Zanthoxylum bungeanum extracts obtained with different solvents, and the result showed that the antifeeding activity of the alcohol extracts was the strongest (93.38%), and the leaf consumption in the treatment and the control showed extremely significant difference (t = 4.23, t_0.01 = 3.25, P = 0.0022), followed by the dichloromethane extracts (47.64%), while the other three solvents (water, acetone, and n-hexane) could not extract the active antifeeding components from Z. bungeanum. The larval consumption of tobacco leaf discs treated with the alcohol extracts of Z. bungeanum and NaCl solution were significantly less than their corresponding controls. The mean larval consumption of the treated leaf discs decreased with ever-increasing dosage, and the consumption of tobacco leaf discs coated with different doses of alcohol extracts of Z. bungeanum or NaCl solution showed extremely significant difference (F_{alcohol extract of Z. bungeanum} = 3.88, F_0.01 = 3.58, P = 0.0064; F_{NaCl solution} = 54.29, F_0.01 = 3.58, P = 0.0000), with maximum antifeeding effects at a dosage of 30 μL per 1.5 cm ID leaf disc. We further tested the larval consumption of tobacco leaf discs treated with alcohol extracts of Z. bungeanum in saturated NaCl solution mixed in different volume ratios, and the result showed that the choice antifeeding percentages of the treatments with 15 μL or more Z. bungeanum alcohol extracts were higher than 90%, among which the mixture with 25:15 volume ratio of Z. bungeanum alcohol extracts and saturated NaCl solution exhibited the strongest antifeeding activity, and the mean consumed leaf area of tobacco leaf discs coated with this blend was only 0.10 mm². In the further test on feeding dose-response of the 25:15 mixture, the mean leaf consumption decreased linearly with ever-increasing dosage, with a regression equation y = ?3.9356x + 120.78(R² = 0.9998), and the 30 μL dose could completely inhibit H. armigera feeding.     

Electrophysiological responses of female Helicoverpa armigera (Hübner) (Lepidoptera; Noctuidae) to synthetic host odours

Studies were conducted to investigate the electroantennographic (EAG) responses of adult female Helicoverpa armigera to a range of known and putative kairomone components. The studies show that at a given dose the EAG responses elicited by a series of straight-chain aliphatic primary alcohols were not dependent on volatility since butan-1-ol and pentan-1-ol elicited EAG responses that were significantly smaller than those elicited by hexan-1-ol. The amplitudes of responses to hexan-1-ol were found to be dose dependent with a dose of 10(-1) μmol at source in a non-volatile solvent eliciting the largest response. Similarly, changes in functionality in a range of C(6) straight-chain aliphatic compounds significantly changed the amplitude of response elicited, with aldehydes eliciting smaller responses than the related primary alcohols and saturated compounds eliciting higher responses than related unsaturated compounds. Of the range of nine host plant-produced terpenoids tested, ocimene and beta-phellandrene elicited the highest responses and of the six aromatic compounds tested phenylacetaldehyde and benzaldehyde elicited the largest responses, at the doses tested. The significance of these findings for analysis of floral odours by gas chromatography linked to electroantennography as a means of identifying kairomone components attractive to H. armigera are discussed.     

Ovicidal activity of Couroupita guianensis (Aubl.) against cotton bollworm Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

There is a great demand for food with increasing population of the world. Many chemical fertilisers and pesticides are used to increase crop productivity. Over use of pesticides and fertilisers has affected the environment and non-target organisms severely; minor pests have become major destructive pests. Hence, eco-friendly approaches are needed. The present study reports the effect of hexane, chloroform and ethyl acetate extract of Couroupita guianensis on the eggs of Helicoverpa armigera. All the extracts showed ovicidal activity, and among them hexane extract showed 64.28% ovicidal activity with LC₅₀ value of 2.62% and regression of (R ²) 83.5%. All the original data showed normality and homogeneity. The active hexane extract was fractionated; finally, eight fractions were obtained and studied at different concentrations. Among the fractions, fraction 8 showed 75% ovicidal activity with R ² of 90%. It showed LC₅₀ value of 322.94?ppm with χ ² value at 0.5%. C. guianensis could be used in pest management programme.     

Variation in susceptibility of Helicoverpa armigera (Hübner) and Helicoverpa punctigera (Wallengren) (Lepidoptera: Noctuidae) in Australia to two Bacillus thuringiensis toxins

Intra-specific variation in susceptibility of Helicoverpa armigera (Hübner) and Helicoverpa punctigera (Wallengren) in Australia to the Cry1Ac and Cry2Ab delta-endotoxins from Bacillus thuringiensis (Berliner) (Bt) was determined to establish a baseline for monitoring changes that might occur with the use of Bt cotton. Strains of H. armigera and H. punctigera were established from populations collected primarily from commercial farms throughout the Australian cotton belts. Strains were evaluated for susceptibility using two bioassay methods (surface treatment and diet incorporation) by measuring the dose response for mortality (LC50) and growth inhibition (IC50). The variation in LC50 among H. armigera (n=17 strains) and H. punctigera (n=12 strains) in response to Cry1Ac was 4.6- and 3.2-fold, respectively. The variation in LC50 among H. armigera (n=19 strains) and H. punctigera (n=12 strains) to Cry2Ab was 6.6- and 3.5-fold, respectively. The range of Cry1Ac induced growth inhibition from the 3rd to 4th instar in H. armigera (n=15 strains) was 3.6-fold and in H. punctigera (n=13 strains) was 2.6-fold, while the range of Cry2Ab induced growth inhibition from neonate to 3rd instar in H. armigera (n=13 strains) was 4.3-fold and in H. punctigera (n=12 strains) was 6.1-fold. Variation in susceptibility was also evaluated for two age classes (neonates and 3rd instars) in laboratory strains of H. armigera and H. punctigera. Neonates of H. punctigera had the same or higher sensitivity to Bt than 3rd instars. Neonates of H. armigera were more sensitive to Cry2Ab than 3rd instars, while being less sensitive to Cry1Ac than 3rd instars. Differences in the two methods of bioassay used affected relative sensitivity of species to Bt toxins, highlighting the need to standardize bioassay protocols.     

Population Genetic Structure of the Cotton Bollworm Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) in India as Inferred from EPIC-PCR DNA Markers

Helicoverpa armigera is an important pest of cotton and other agricultural crops in the Old World. Its wide host range, high mobility and fecundity, and the ability to adapt and develop resistance against all common groups of insecticides used for its management have exacerbated its pest status. An understanding of the population genetic structure in H. armigera under Indian agricultural conditions will help ascertain gene flow patterns across different agricultural zones. This study inferred the population genetic structure of Indian H. armigera using five Exon-Primed Intron-Crossing (EPIC)-PCR markers. Nested alternative EPIC markers detected moderate null allele frequencies (4.3% to 9.4%) in loci used to infer population genetic structure but the apparently genome-wide heterozygote deficit suggests in-breeding or a Wahlund effect rather than a null allele effect. Population genetic analysis of the 26 populations suggested significant genetic differentiation within India but especially in cotton-feeding populations in the 2006–07 cropping season. In contrast, overall pair-wise F _ST estimates from populations feeding on food crops indicated no significant population substructure irrespective of cropping seasons. A Baysian cluster analysis was used to assign the genetic make-up of individuals to likely membership of population clusters. Some evidence was found for four major clusters with individuals in two populations from cotton in one year (from two populations in northern India) showing especially high homogeneity. Taken as a whole, this study found evidence of population substructure at host crop, temporal and spatial levels in Indian H. armigera, without, however, a clear biological rationale for these structures being evident.     

Effects of larval host plants on over-wintering preparedness and survival of the cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

Laboratory colonies of cotton bollworm larvae, Helicoverpa armigera, kept at 20 degrees C under a photoperiod of L:D=10:14 were fed on five host plants (cotton, corn, kidney bean, tobacco and tomato) and an artificial diet (control) to determine the effects of larval host quality on survival and pupal over-wintering preparedness. A separate experiment showed that diapausing pupae weighed more and contained greater nutrient stores than did non-diapausing pupae. Diapausing pupae reared on different host plants showed significant differences in terms of over-wintering reserve storage, and degree of cold-hardiness (extent of low-molecular-weight substances and SCPs), and survivorship. The more nutrients the host plant had, the more the pupae weighed and the higher the levels of total lipids and glycogen. Body water content was also significantly affected by larval food quality. The mean pupal super-cooling capacities varied significantly from -16.7 to -18.9 degrees C according to host plants the larvae feed on, and these significantly related to water content, pupal weight, lipid and glycogen content, and the levels of glycerol. Levels of trehalose, glycerol, and inositol, which were mainly low-molecular-weight substances, showed no significant differences among different host plants, except for trehalose. Pupal mortality varied from 39.7% on corn to 3.3% on the artificial diet, which was significantly related to pupal weight, total lipid content, trehalose levels, and super-cooling points. These results suggest that larval food quality can affect survival and influence the over-wintering preparedness of the cotton bollworm.     

Effects of larval host plants on over-wintering physiological dynamics and survival of the cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

Laboratory colonies of cotton bollworm larvae, Helicoverpa armigera, were kept at 20 °C under a photoperiod of L:D = 10:14 and fed on three host plants (cotton, tobacco, kidney bean) and an artificial diet (control) to determine the dynamic effects of larval host quality on over-wintering physiology and mortality. Energy reserves (glycogen and lipid), super-cooling points (SCPs), low-molecular-weight sugars, temperature, and mortality were monitored from November 2002 to April 2003. Lipid content did not change much for each group during over-wintering, but differed according to larval host plants. Larval host plants obviously influence the amount of glycogen, as does time of year: glycogen was lowest in February and increased in early spring. During winter, the mean pupal SCPs increased the most in February, then decreased, and were also affected by larval host plant, i.e. over-wintering pupae reared on kidney bean had the highest SCPs. Levels of glycerol and inositol differed significantly among host plants and months, which peaked in February. Pupal mortality also varied according to larval host plants and time: pupae reached their highest mortality in March and showed host plant differences in January. Records show that February was the coldest month during the period we observed, which corresponded closely to changes in over-wintering characteristics.     

Last instar larva and pupa of Melipotis cellaris (Guenée) (Lepidoptera: Noctuidae)

The last instar larva and pupa of Melipotis cellaris (Guenée) are described and illustrated, based on specimens collected in northern Chile, associated with Acacia macracantha (Fabaceae).     

Molecular cloning and bacterial expression of pheromone binding protein in the antennae of Helicoverpa armigera (Hübner)

A cDNA clone coding for pheromone binding protein was isolated from the antennae of Helicoverpa armigera by RT-PCR and (5'/3')-RACE technique. The full-length of H. armigera pheromone binding protein (HarmPBP) was 952 bp, possessing 162 amino acid residues including a signal peptide of 20 amino acids. Its predicted molecular weight and isoelectric point were 18.26 kDa and 5.23, respectively. This deduced amino acid sequence shared some common structural features with odorant-binding proteins from several moth species, including the six conserved cysteine motif, a typical characteristic of insect's odorant-binding proteins. Northern blot showed that HarmPBP is specifically expressed in the antennae of Helicoverpa armigera and more abundantly expressed in male than female. During the antennal development, HarmPBP is first expressed about 4 days prior to adult eclosion and rises to a plateau 2 days prior to adult eclosion. In order to obtain sufficient PBP for further determining its biochemical and physiological properties, a bacterical expression vector of PBP was constructed and successfully expressed in Escherichia coli. The recombinant PBP was shown to cross-react with an anti-PBP antiserum from Antheraea polyphemus. Polyclonal antibodies against HarmPBP were used to mark the distribution of the protein in olfactory sensilla. Very strong labeling was observed in the sensillum lymph of the hair lumen and of the sensillum-lymph cavity. In the male, HarmPBP is expressed in sensilla trichodea and not in sensilla basiconica, while in the female, it is expressed both in sensilla basiconica and sensilla trichodea.     

Enhancement of attraction to sex pheromone of Grapholita molesta (Busck) (Lepidoptera: Tortricidae) by structurally unrelated sex pheromone compounds of Conogethes punctiferalis (Guenée) (Lepidoptera: Crambidae)

Grapholita molesta (Busck) (the oriental fruit moth; OFM) and Conogethes punctiferalis (Guenée) are both fruit and stem borers with overlapping habitats, occurrences, and outbreak cycles. These two species are in different families and they have completely different sex pheromone components. Here, the effects of the sex pheromone components of C. punctiferalis, (E)-10-hexadecenal (E10-16:Ald) and (Z)-10-hexadecenal (Z10-16:Ald) and their mixture, were evaluated on the sexual communication of OFM by examining electrophysiological (EAG) and behavioral responses. We found that a considerably large amount of E10-16:Ald or Z10-16:Ald and a tiny amount of OFM pheromone elicited comparable EAG responses in OFM males, indicating the low antennal olfactory sensitivity of OFM males to the heterospecific pheromone compounds. In two different peach orchard parcels, captures of OFM by their pheromone lures baited in delta traps were increased by at least 1.5 times when OFM pheromone lures were combined with lures that contained 1000 μg of either E10-16:Ald, Z10-16:Ald or their mixture. In two other pear orchard parcels, both E10-16:Ald and Z10-16:Ald increased captures of OFM in Unitraps in a dose-dependent manner with more than a four-fold increase. Therefore, the enhanced attractiveness of OFM lures by the two interspecific pheromones suggests their potential application to improve mass trapping, population monitoring and mating disruption of OFM.     

Possible mechanisms mitigating reproductive interference supposed between Xylena fumosa (Butler) and X. formosa (Butler) (Lepidoptera: Noctuidae: Xyleninae), both by-captured in a commercial pheromone trap for Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae: Heliothinae)

As differences in sex pheromones usually act as a barrier against interspecific mating, the fact that the closely related moths Xylena fumosa (Butler) and X. formosa (Butler) (Lepidoptera: Noctuidae: Xyleninae) are commonly attracted to the sex pheromone lure for Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae: Heliothinae) seems unlikely, suggesting that some mechanisms exist between them to mitigate a potential reproductive interference in pheromone communications. Thus, we examined the overlaps in ecological niches of the two species, and their attractiveness to commercial pheromone lures for other moths, through pheromone trap surveys and published reports. Long-term field surveys using traps baited with five to six types of artificial sex pheromone lures including those designed for H. armigera, and information in published works revealed that there was a nonnegligible overlap between the two species in terms of seasonal timing of captures, geographic habitats and host plants. These factors, therefore, did not appear to provide conclusive evidence mitigating reproductive interference. However, there was a slight but significant difference between them in the trends of attraction to different artificial pheromone lures; X. formosa was attracted albeit in a small number to the lures for Plutella xylostella (Linnaeus) (Lepidoptera: Plutellidae), whereas X. fumosa was only attracted to those for H. armigera. This suggests that differences in pheromone composition, rather than differences in temporal and spatial distributions, between the two species could play a more significant role in premating reproductive isolations.     

Biological aspects of Autoplusia egena (Guenée) (Lepidoptera: Noctuidae, Plusiinae)

Autoplusia egena (Guenée) is a moth distributed throughout the Western hemisphere. The larvae have already been found feeding on several different plant families, including important crops such soybeans and beans. To contribute to the knowledge of its biology in laboratory conditions, and considering the duration and the morphometry of each development stage, a laboratorial rearing was accomplished at 25 +/- 1 degrees C, 70 +/- 10% RH and 14h photophase. The larvae were fed with black bean leaves and the adults were fed with a honey solution at 10%. The results, expressed by the mean (+/- SE), for the periods of each stage were: egg 3.0 +/- 0.00 d; larva 15.7 +/- 1.25 d; pre-pupa 1.9 +/- 0.05 d; pupa 8.8 +/- 0.09 d and adult: longevity 12.1 +/- 0.95 d, pre-oviposition 5.4 +/- 0.50 d, oviposition 6.3 +/- 1.10 d and post-oviposition 1.4 +/- 0.87 d. The larvae went through five instars, for which the mean width of the cephalic capsules were 0.302 +/- 0.001 mm; 0.500 +/- 0.003 mm; 0.854 +/- 0.008 mm; 1.424 +/- 0.011 mm and 2.744 +/- 0.053 mm. The average length, width, and weight of the pupae were 16.965 +/- 0.003 mm, 4.674 +/- 0.040 mm and 0.217 +/- 0.003 g, respectively.     

Organophosphorus insecticides synergize pyrethroids in the resistant strain of cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) from West Africa

Helicoverpa armigera (Hübner) populations from West Africa recently developed resistance to pyrethroid insecticides through enhanced metabolism by mixed-function oxidases. The combination index method was used to study the synergism of pyrethroids by organophosphorus insecticides. Several mixtures of insecticides currently registered to control cotton pest complex in West Africa were tested, including: cypermethrin/ethion, cypermethrin/profenofos, deltamethrin/ triazophos, deltamethrin/chlorpyriphos, cyfluthrin/chlorpyriphos, and betacyfluthrin/chlorpyriphos. In the resistant strain, the organophosphorus insecticides significantly increased the toxicity of pyrethroids suppressing the resistance effect, either by additive or synergistic effects. Significant synergism was shown for the following mixtures: cypermethrin/ethion, deltamethrin/triazophos, and deltamethrin/chlorpyriphos. The use of synergism from these insecticide mixtures should prove to be an additional tool in the overall resistance management strategy because the pyrethroid resistance in H. armigera from West Africa is not yet stable, decreasing between cotton seasons and increasing with treatments. In absence of selection, the susceptibility of H. armigera to insecticides should be restored.     

A comparison of artificial diet and hybrid sweet corn for the rearing of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) based on life table characteristics

The demographic characteristics of Helicoverpa armigera (Hübner) reared on hybrid sweet corn (Zea mays L. variety saccharata) (hybrid super sweet corn KY bright jean) and on an artificial diet were compared by using the age-stage, two-sex life table. Because the hatch rate of eggs varies with maternal age, age-specific fecundity was calculated based on the numbers of hatched eggs to reveal the biological characteristics of H. armigera accurately. The intrinsic rate of increase (r), finite rate (λ) and mean generation time (T) of H. armigera were 0.0853 d(-1), 1.0890 d(-1), and 46.6 d, respectively, on Z. mays and 0.1015 d(-1), 1.1068 d(-1), and 46.3 d, respectively, on the artificial diet. There were significant differences in the intrinsic rate of increase and finite rate between two treatments. The age-stage life expectancy and reproductive value also were calculated. The relationships among the net reproductive rate, the mean female fecundity, the number of emerged females, and the total number of individuals used in the life table study are consistent with theoretical expectations. We recommend the age-stage, two-sex life table for use in insect demographic studies to incorporate both sexes and the variation in developmental rate among individuals and to obtain accurate population parameters. The artificial diet is more suitable for the mass rearing of H. armigera.