Feeding behavior of bollworm and tobacco budworm (Lepidoptera: Noctuidae) larvae in mixed stands of nontransgenic and transgenic cotton expressing an insecticidal protein

Feeding behavior of third-instar bollworm, Helicoverpa zea (Boddie), and tobacco budworm, Heliothis virescens (F.), was observed in pure and mixed stands of nontransgenic and transgenic cotton (BTK), Gossypium hirsutum L., expressing an insecticidal protein CryIA(c) from a bacterium, Bacillus thuringiensis Berliner subsp. kurstaki. Five plant stands composed of BTK and non-BTK plants were evaluated; two pure stands and three mixed stands. Percentage ratios of BTK to non-BTK plants in the stands were 100:0, 75:25, 50:50, 25:75 and 0:100, respectively. In all stands with BTK plants, fewer bollworm and tobacco budworm larvae were found on BTK plants than non-BTK plants 24 h after infestation with third instars. At 48 h, significantly fewer tobacco budworm larvae, but not fewer bollworm larvae, were found on BTK plants. However, the number of larvae of either insect did not increase on non-BTK plants compared with the initial infestation density of three larvae per plant. The number of obacco budworm injured flower buds, and capsules was lower in all plant stands containing BTK plants compared with the pure stand of non-BTK at 48 h after infestation. Higher numbers of larvae on non-BTK plants were possibly the result of larval intoxication, reduced feeding, and increased plant abandonment and death on BTK plants rather than a classical feeding preference. Unexpectedly, the number of flower buds and capsules injured by bollworm and tobacco budworm when averaged per plant for all plants in a stand, differed little among the 75:25, 50:50 and 25:75 plant mixtures. These data suggest that larvae of both species frequently moved among plants, feeding indiscriminately on BTK and non-BTK plants.     

Effects of cotton cultivar on fitness costs associated with resistance of pink bollworm (Lepidoptera: Gelechiidae) to Bt cotton

Fitness costs associated with insect resistance to transgenic crops producing toxins from Bacillus thuringiensis (Bt) reduce the fitness on non-Bt refuge plants of resistant individuals relative to susceptible individuals. Because costs may vary among host plants, choosing refuge cultivars that increase the dominance or magnitude of costs could help to delay resistance. Specifically, cultivars with high concentrations of toxic phytochemicals could magnify costs. To test this hypothesis, we compared life history traits of three independent sets of pink bollworm, Pectinophora gossypiella (Saunders), populations on two cotton cultivars that differed in antibiosis against this cotton pest. Each set had an unselected susceptible population, a resistant population derived by selection from the susceptible population, and the F1 progeny of the susceptible and resistant populations. Confirming previous findings with pink bollworm feeding on cotton, costs primarily affected survival and were recessive on both cultivars. The magnitude of the survival cost did not differ between cultivars. Although the experimental results did not reveal differences between cultivars in the magnitude or dominance of costs, modeling results suggest that differences between cultivars in pink bollworm survival could affect resistance evolution. Thus, knowledge of the interaction between host plants and fitness costs associated with resistance to Bt crops could be helpful in guiding the choice of refuge cultivars.     

Overwintering cost associated with resistance to transgenic cotton in the pink bollworm (Lepidoptera: Gelechiidae)

Fitness costs associated with resistance to transgenic crops producing toxins from Bacillus thuringiensis (Bt) may have important effects on the evolution of resistance. We investigated overwintering costs in pink bollworm, Pectinophora gosypiella (Saunders), strains with different degrees of resistance to Bt cotton. Frequency of resistant individuals in a strain was not associated with induction of diapause or emergence from diapause in early winter. Emergence from diapause in the spring was 71% lower in three highly resistant strains than in two heterogeneous strains from which the resistant strains were derived. This underestimates the overwintering cost because the frequency of the resistance allele was relatively high in the heterogeneous strains. Emergence in the spring in hybrid progeny from crosses between the resistant and heterogeneous strains was greater than in resistant strains but did not differ from susceptible strains, showing that the overwintering cost was recessive to some extent.     

Behavior of bollworm (Lepidoptera: Noctuidae) larvae on genetically engineered cotton

Reports of bollworm, Helicoverpa zea (Boddie), larvae feeding in white flowers of Bollgard cotton have been relatively common since the commercialization of this technology in 1996. Field studies were conducted in Louisiana to determine if differences in bollworm larval behavior occuron non-Bollgard (cultivar 'Deltapine 5415') and Bacillus thuringiensis (Bt), Bollgard ('NuCOTN 33B') cottons. Larvae were placed on the terminal foliage of either single cotton plants or on all plants within 1-m row micro-plots. On preflowering cotton plants, significantly more bollworms moved from the site of infestation (terminal) on Bollgard plants compared with that on non-Bollgard plants. On individual flowering plants, the number of nodes larvae moved from the terminal and the number of infested bolls was greater on Bollgard cotton plants. Similar differences between Bollgard and non-Bollgard plants in the percentage of infested terminals and squares were observed at 48-h after infestation when 1-m rows were infested. These data will be used to refine scouting protocols for bollworm larvae on Bollgard cotton.     

Antennal response of cotton bollworm (Heliocoverpa armigera) to volatiles in transgenic Bt cotton

Abstract:  Volatile compositions in transgenic Bt cotton variety 'GK-97' and its parental variety 'Simian No. 3' were analysed, and antennal response of cotton bollworm ( Heliocoverpa armigera ) to the volatiles were investigated. Volatile components in Bt cotton and regular cotton were almost same, except for absence of a sesquiterpene and a minor unknown compound in regular cotton. The results of gas chromatography-electroantennographic detection (GC-EAD) experiments showed that cotton bollworm antennae responded to seven compounds and two minor unknown compounds in Bt volatiles, among which concentrations of α -pinene and β -pinene in Bt cotton were relatively higher than those in regular cotton, and one minor unknown compound was absent in regular cotton.     

Plant-toxin interactions in transgenic Bt cotton and their effect on mortality of Helicoverpa armigera (Lepidoptera: Noctuidae)

In Australia, transgenic cotton plants expressing the cry1Ac gene from Bacillus thuringiensis Berliner variety kurstaki are less toxic to first-instar Helicoverpa armigera (Hübner) after the plant is producing fruit. We developed two bioassay methods (leaf mush, leaf disk) to test if the physiological state of the plants explained changes in toxicity and a third method (diet incorporation) was developed to quantify the toxicity of Bt leaves when mixed in chickpea diet. Cry1Ac protein was less toxic to H. armigera larvae when the protein was mixed with leaves from fruiting versus presquare conventional cotton. Differences in LC50 varied from 2.4- to 726-fold, depending on the source of toxin and conventional plant material. These results suggest that plant-toxin interactions in fruiting cotton are reducing the toxicity of the Cry1Ac protein. The possible role of tannins in these changes is discussed.     

Mitochondrial genome of the cotton bollworm Helicoverpa armigera (Lepidoptera: Noctuidae) and comparison with other Lepidopterans

We present the complete sequence of the mitochondrial genome (mitogenome) of the cotton bollworm Helicoverpa armigera. The 15,347-bp mitogenome of H. armigera was arranged in the same order described for all other sequenced lepidopterans, which differs from the most common type found in insects, due to the movement of trnM to a position 5'-upstream of trnI. The gene overlap in the H. armigera mitogenome is totally 23 bp in six locations. The H. armigera mitogenome has a total of 175 bp of intergenic spacer sequences spread over 14 regions ranging in size from 1 to 45 bp. The nucleotide composition of the whole mitogenome of H. armigera is highly A+T biased, accounting for 80.97%, with a slightly positive AT skewness and negative GC skewness, indicating the occurrence of more A than T, C more than G. The protein-encoding genes have typical mitochondrial start codons, except for cox1, which contains the unusual CGA. The cox1, cox2, and nad4 genes have incomplete stop codons (T). The lrRNA and srRNA genes are 1395 and 794-bp long, respectively. All tRNAs have a typical cloverleaf structure of mitochondrial tRNAs, except for trnS1(AGN), the dihydrouridine arm of which could not form a stable stem-loop structure. The H. armigera A+T-rich region contains a conserved structure combining the motif ATAGA and a 19-bp poly-T stretch, but absence of the 9-bp poly-A element upstream of trnM.     

Fitness costs and maternal effects associated with resistance to transgenic cotton in the pink bollworm (Lepidoptera: Gelechiidae).

Transgenic cotton producing a Bacillus thuringiensis (Bt) toxin is widely used for controlling the pink bollworm, Perctinophora gossypiella (Saunders). We compared performance of pink bollworm strains resistant to Bt cotton with performance of their susceptible counterparts on non-Bt cotton. We found fitness costs that reduced survival on non-Bt cotton by an average of 51.5% in two resistant strains relative to the susceptible strains. The survival cost was recessive in one set of crosses between a resistant strain and the susceptible strain from which it was derived. However, crosses involving an unrelated resistant and susceptible strain indicated that the survival cost could be dominant. Development time on non-Bt cotton did not differ between the two related resistant and susceptible strains. A slight recessive cost affecting development time was suggested by comparison of the unrelated resistant and susceptible strains. Maternal effects transmitted by parents that had eaten Bt-treated artificial diet as larvae had negative effects on embryogenesis, adult fertility, or both, and reduced the ability of neonates to enter cotton bolls. These results provide further evidence that fitness costs associated with the evolution of resistance to Bt cotton are substantial in the pink bollworm.     

Cytochrome P450 monooxygenases in the cotton bollworm (Lepidoptera: Noctuidae): tissue difference and induction

Differences in the microsomal P450 monooxygenase system and its inducibility by pentamethylbenzene (PMB) and naphathalene (NA) were investigated in midgut and fatbody tissues of the cotton bollworm, Helicoverpa armigera (Boddie), larvae. Orthogonal array design was used to establish the optimal conditions for measuring Aldrin epoxidation (AE). The optima for AE were similar for the midgut and the fatbody at a temperature of 30 degrees C, pH 7.4, and a time of 10 min. In comparison to fatbody, the midgut had higher levels of total cytochrome P450s, p-nitroanisole O-demethylation (ODM) and AE. In vivo administration of 0.2% PMB or 0.2% NA resulted in higher microsomal protein content and levels of total cytochrome P450 as well as the two examined monooxygenase activities. Total cytochrome P450 and ODM activity were induced to a greater degree in the fatbody. In the midgut, PMB was significantly more effective on ODM than NA. Differences existed in SDS-PAGE profiles between the midgut and the fatbody. The induction of the midgut with PMB and of the fatbody with NA and PMB resulted in marked intensification of the protein bands with molecular masses of 59,100, 53,400, 50,400 Da.     

Gene-flow between populations of cotton bollworm Helicoverpa armigera (Lepidoptera: Noctuidae) is highly variable between years

Both large and small scale migrations of Helicoverpa armigera Hübner in Australia were investigated using AMOVA analysis and genetic assignment tests. Five microsatellite loci were screened across 3142 individuals from 16 localities in eight major cotton and grain growing regions within Australia, over a 38-month period (November 1999 to January 2003). From November 1999 to March 2001 relatively low levels of migration were characterized between growing regions. Substantially higher than average gene-flow rates and limited differentiation between cropping regions characterized the period from April 2001 to March 2002. A reduced migration rate in the year from April 2002 to March 2003 resulted in significant genetic structuring between cropping regions. This differentiation was established within two or three generations. Genetic drift alone is unlikely to drive genetic differentiation over such a small number of generations, unless it is accompanied by extreme bottlenecks and/or selection. Helicoverpa armigera in Australia demonstrated isolation by distance, so immigration into cropping regions is more likely to come from nearby regions than from afar. This effect was most pronounced in years with limited migration. However, there is evidence of long distance dispersal events in periods of high migration (April 2001-March 2002). The implications of highly variable migration patterns for resistance management are considered.     

Boll injury and yield losses in cotton associated with brown stink bug (Heteroptera: Pentatomidae) during flowering

Brown stink bug, Euschistus servus (Say), was infested on cotton, Gossypium hirsutum L., plants during reproductive stages to determine the effects on boll injury and seedcotton yield. During each week in 2002 and 2003, significantly more bolls with > or = 1 injured locule, bolls with > or = 2 injured locules, and bolls with discolored lint were recorded on stink bug-infested plants compared with that on noninfested plants. Significantly fewer bolls displayed external injury on the boll exocarp compared with bolls with only internal locule injury. Boll injury was significantly underestimated by the presence of external symptomology. The boll population increased 6.6- and 5.1-fold from weeks 1-5 in 2002 and 2003, respectively. There was a corresponding 6.2- and 4.6-fold increase in 2002 and 2003, respectively, for total bolls injured from weeks 1-5. Percentage of boll injury ranged from 10.7 (week 4) to 27.4 (week 2) in 2002 and from 9.2 (week 3) to 16.0 (week 2) in 2003. Percentage of injury was greatest during weeks 1 and 2 in both years and also in week 5 in 2002. Brown stink bug significantly reduced seedcotton yield of bolls present on cotton plants during weeks 1, 2, and 5 in 2002 and in weeks 4 and 5 in 2003. However, total seedcotton yield, as a function of bolls exposed to brown stink bug and subsequent bolls produced on plant in the absence of stink bugs, was not significantly different for plots infested during weeks 1-4 in 2002 and weeks 1-3 in 2003. Flowering period and boll population influence the severity of stink bug injury on seedcotton yield. Infestation timing and number of bolls should be considered, in addition to insect densities, when initiating treatments against brown stink bug.     

Early detection of field-evolved resistance to Bt cotton in China: cotton bollworm and pink bollworm

Transgenic crops producing Bacillus thuringiensis (Bt) toxins kill some major insect pests, but pests can evolve resistance and thereby reduce the effectiveness of such Bt crops. The main approach for slowing pest adaptation to Bt crops uses non-Bt host plants as "refuges" to increase survival of susceptible pests. To delay evolution of pest resistance to cotton producing Bt toxin Cry1Ac, several countries have required refuges of non-Bt cotton, while farmers in China have relied on "natural" refuges of non-Bt host plants other than cotton. This strategy is designed for cotton bollworm (Helicoverpa armigera), which attacks many crops and is the primary target of Bt cotton in China, but it does not apply to pink bollworm (Pectinophora gossypiella), which feeds almost entirely on cotton in China. Here we review evidence of field-evolved resistance to Cry1Ac by cotton bollworm in northern China and by pink bollworm in the Yangtze River Valley of China. For both pests, results of laboratory diet bioassays reveal significantly decreased susceptibility of field populations to Cry1Ac, yet field control failures of Bt cotton have not been reported. The early detection of resistance summarized here may spur countermeasures such as planting Bt cotton that produces two or more distinct toxins, increased planting of non-Bt cotton, and integration of other management tactics together with Bt cotton.     

Correlating differences in larval survival and development of bollworm (Lepidoptera: Noctuidae) and fall armyworm (Lepidoptera: Noctuidae) to differential expression of Cry1A(c) delta-endotoxin in various plant parts among commercial cultivars of transgenic Bacillus thuringiensis cotton

Differences in larval survival and development of bollworm, Helicoverpa zea (Boddie), and fall armyworm, Spodoptera frugiperda (J. E. Smith), respectively, were found to exist among commercially available Cry1A(c) transgenic Bacillus thuringiensis Berliner (Bt) varieties. Using a quantification assay (ELISA) to measure the levels of delta-endotoxin in two of these varieties ('DP 451B/RR' and 'NuCOTN 33B'), differences in the amount of delta-endotoxin present in various plant parts was correlated with larval survival of bollworms and larval development of fall armyworms throughout the growing season. Larvae that were fed on DP 451B/RR completed development faster and exhibited better survivorship than those larvae fed NuCOTN 33B, whereas lower levels of delta-endotoxin were generally detected in plant parts from DP 451B/RR compared with NuCOTN 33B. These differences may impact population dynamics of these pests which may be a critical factor in managing resistance to Bt. Furthermore, the utility of using this system for providing information to the grower concerning varietal choices may be more common in the future.     

Inheritance of Cry1Ac resistance and associated biological traits in the cotton bollworm, Helicoverpa armigera (Lepidoptera: Noctuidae)

The analysis of reciprocal genetic crosses between resistant Helicoverpa armigera strain (BH-R) (227.9-fold) with susceptible Vadodara (VA-S) strain showed dominance (h) of 0.65-0.89 and degree of dominance (D) of 0.299-0.782 suggesting Cry1Ac resistance as a semi-dominant trait. The D and h values of F₁ hybrids of female resistant parent were higher than female susceptible parent, showing maternally enhanced dominance of Cry1Ac resistance. The progeny of F₂ crosses, backcrosses of F₁ hybrid with resistant BH-R parent did not differ significantly in respect of mortality response with resistant parent except for backcross with female BH-R and male of F₁ (BH-R × VA-S) cross, suggesting dominant inheritance of Cry1Ac resistance. Evaluation of some biological attributes showed that larval and pupal periods of progenies of reciprocal F₁ crosses, backcrosses and F₂ crosses were either at par with resistant parent or lower than susceptible parent on treated diet (0.01 μg/g). The susceptible strain performed better in terms of pupation and adult formation than the resistant strain on untreated diet. In many backcrosses and F₂ crosses, Cry1Ac resistance favored emergence of more females than males on untreated diet. The normal larval period and the body weight (normal larval growth) were the dominant traits associated with susceptible strain as contrast to longer larval period and the lower body weight (slow growth) associated with resistance trait. Further, inheritance of larval period in F₂ and backcross progeny suggested existence of a major resistant gene or a set of tightly linked loci associated with Cry1Ac sensitivity.     

Sensitivity analysis of a spatially-explicit stochastic simulation model of the evolution of resistance in Helicoverpa zea (Lepidoptera: Noctuidae) to Bt transgenic corn and cotton

The sensitivities of a model simulating the evolution of resistance in Helicoverpa zea to Bt toxins in transgenic crops were investigated by examining effects of each of the model parameters on the frequency of resistance alleles after 8 yr. The functional dominance of resistance alleles and the initial frequency of those alleles had a major impact on resistance evolution. The survival of susceptible insects on the transgenic crops and the population dynamics of the insect, driven by winter survival and reproductive rates, were also important. In addition, agricultural practices including the proportion of the acreage planted to corn, and the larval threshold for spraying cotton fields affected the R-allele frequency. Many of these important parameters are inherently variable or cannot be measured with accuracy, so model output cannot be interpreted as being a forecast. However, this analysis is useful in focusing empirical research on those aspects of the insects' life system that have the largest effects on resistance development, and indicates ways in which to improve products and agricultural practices to increase the expected time to resistance. The model can thus be used as a scientific basis for devising a robust resistance management strategy for Bt crops.     

Cannibalism of Helicoverpa zea (Lepidoptera: Noctuidae) from Bacillus thuringiensis (Bt) transgenic corn versus non-Bt corn

Because of the importance of cannibalism in population regulation of Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) in corn, Zea mays L., it is useful to understand the interactions between Bacillus thuringiensis (Bt) transgenic corn and cannibalism. To determine the effects of Bt corn on cannibalism in H. zea, pairs of the same or different instars were taken from Bt or non-Bt corn and placed on artificial diet in proximity. Cannibalism occurred in 91% of pairs and was approximately 7% greater for pairs of larvae reared from Bt transgenic corn (95%) than from non-Bt corn (88%). Also, first instar by first instar pairs had a lower rate of cannibalism than other pairs. Time until cannibalism was not different for larvae from Bt corn versus non-Bt corn. Pupation rate of cannibals and surviving victims was not different for pairs from Bt corn versus non-Bt corn. Finally, cannibalism increased pupation rate of cannibals from both Bt and non-Bt corn by approximately 23 and 12%, respectively, although the increases were not significant. Thus, negative effects of Bt on larvae were compensated by increased cannibalism in comparison with larvae reared on non-Bt corn, which increased larval survival to levels comparable with larvae reared on non-Bt plants.     

磁场对棉铃虫生长发育及繁殖的影响

棉铃虫是华北地区重要的农业害虫,磁场可能对该害虫的生长发育存在显著影响.本试验设置不同磁场强度(零磁场、高磁场和自然磁场),观察棉铃虫幼虫生长发育以及成虫繁殖情况.结果表明:零磁场能明显延长幼虫历期,平均延迟0.4～1.7 d;高磁场对幼虫历期没有明显影响;幼虫存活率、蛹重、羽化率、成虫产卵量及卵孵化率等指标受磁场影响并不显著;表明零磁场延缓幼虫发育,但对其繁殖没有明显影响;由于试验中高磁场强度仅有4 Gs,是否更高磁场有影响还需进一步验证.     

An iridescent virus of the bollworm Heliothis zea (Lepidoptera:Noctuidae)

Larvae of Heliothis zea exhibiting iridescent lavender-blue, blue, blue-green color were obtained in spring field collections in host plants on a road right-of-way in Bolivar County, Mississippi. An iridescent virus was isolated and purified from these larvae. Size range of the virus was 131 to 160 nm, with a mean ± SD of 145 ± 6.5. The nucleic acid content was: DNA, 13.97 ± 1.58% (0.139 μg of DNA/mg of viral protein); RNA, none. Experimental infection was not achieved per os, but the virus was highly virulent by intrahemocoelic injection. The virus from larvae killed in this way was of the same size and morphology as that obtained in the initial isolation.     

Mating compatibility among helicoverpa armigera (Lepidoptera: Noctuidae) occurring on selected host plants and Bt cotton survivors

Helicoverpa armigera (Hiibner) (Lepidoptera: Noctuidae) is a well-known polyphagous insect pest. Mating compatibility among the insects occurring on different host plants is essential for free gene flow among populations. We tested the extent of crossability and fecundity of the insects that survived on Bacillus thuringiensis (Bt) cotton with those occurring on pigeon pea, Cajanus cajun (L.) Millsp., non-Bt cotton, Gossypium hirsutum L.; sunflower, Helianthus annuus; sorghum, Sorghum bicolor L. Moench.; okra, Abelmoschus moschatus Medikus; chickpea, Cicer arietinum L.; marigold, Tagetes spp.; and tomato, Lycopersicum esculentum L., crops. The insects from different crops were freely crossable with those collected from Bt cotton and among themselves. The average fecundity across different crosses ranged from 314.1 to 426.3 in direct and from 305.8 to 421.7 eggs per female in reciprocal crosses. In any given cross, a minimum of 85.89% egg hatch was recorded. Furthermore, the F1 individuals of different cross combinations were found to cross freely with their parents (BC1) and among themselves with similar fecundity and egg hatch. High crossability among H. armigera occurring on different host plants suggests that crop mosaics that may exist in countries such as India could play an important role as natural, nonstructured refugia and prolong the durability of the genes deployed for controlling this insect.     

Estimated frequency of nonrecessive Bt resistance genes in bollworm,Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) in eastern North Carolina

In summer 2000, adult female bollworm moths, Helicoverpa zea (Boddie), were collected from light-traps at four locations near the Tidewater Research Station, Plymouth, NC. Female moths were allowed to lay eggs, and at hatch, 72 larvae from each female were screened for growth rate on normal artificial diet and on diets containing 5.0 microg of either Cry1Ac or Cry2Aa Bt toxin per milliliter of diet. The growth rate bioassays were performed to isolate nonrecessive Bt resistance genes present in field populations of bollworm. We found one individual out of 583 screened that appeared to carry a major gene for resistance to Cry1Ac. Assuming four alleles per individual, the gene frequency is 1/2332 or 0.0003. Other females appeared to have minor genes for Cry1Ac resistance or major genes with lower levels of dominance. We also found one individual out of 646 screened that appeared to carry a major gene for resistance to Cry2Aa. The gene frequency for Cry2Aa resistance was estimated at 1/2584 or 0.00039. Again, other females seemed to carry additional minor resistance genes. Along with other results that indicate partially dominant inheritance of Cry1Ac resistance in bollworm, these allele frequency estimates are important for determining the rate of resistance evolution in H. zea to specific Bt toxins.