Is resistance to Bacillus thuringiensis endotoxin Cry1Ac associated with a change in the behavior of light brown apple moth larvae (Lepidoptera: Tortricidae)?

Bacillus thuringiensis (Bt)-resistant light brown apple moth, Epiphyas postvittana (Walker) (Lepidoptera: Tortricidae), created by selection of a laboratory colony with artificial diets containing the Bt endotoxin Cry1Ac, were used to explore relationships between larval behavior and resistance to toxins. Our hypothesis was that behavioral responses during the first days of exposure to diet are directly related to the toxicity of the diet, as measured by subsequent mortality. We tested two predictions from this hypothesis. The first prediction was that susceptible larvae and resistant larvae exhibit similar behavior on diet without toxins, settling at feeding sites within a few hours. The second prediction was that susceptible and resistant larvae differ in their behavior on Cry1Ac diet to the same degree that their mortality differs, i.e., on Cry1Ac diet, resistant larvae exhibit anorexia and walking to a lesser degree than susceptible larvae. Predictions were tested by making observations over 2 wk, with each larva held individually in a 10-cm-long cylindrical glass arena with two aliquots of diet. The two aliquots consisted of either the same diet (two no-choice treatments: control/control or Cry 1Ac/Cry1Ac) or different diets (one choice treatment: control/Cry 1Ac). The two predictions did not accurately describe larval behavior. On control diet, behavior differed, with resistant larvae settling more quickly than susceptible larvae. On Cry1Ac diet, behavior was more similar than expected. Thus, even though the Bt diet was much less toxic to resistant larvae, resistant larvae seemed to match the ability of susceptible larvae to reduce exposure to Bt diet while increasing exposure to nontoxic control diet.     

Reproductive compatibility of several East and West African Cotesia sesamiae (Hymenoptera: Braconidae) populations and their crosses and backcrosses using Sesamia calamistis (Lepidoptera: Noctuidae) as the host

The relative importance of the braconid Cotesia sesamiae, a gregarious larval parasitoid of lepidopteran stemborers, varies greatly with region in Africa; while the most common parasitoid of noctuid stemborers in eastern Africa, it is rare in western Africa. Thus, several strains of C. sesamiae from Kenya are envisaged for introduction into western Africa. The present study investigates the reproductive compatibility between four populations of C. sesamiae from West Africa and Kenya with the noctuid Sesamia calamistis as the host using reciprocal crosses as well as backcrosses of hybrid females with males of the parental populations. Searching time of the male for the female and mating period varied significantly with couple and ranged between 0.78-1.9 min and 3.4-12.8 s, respectively. Crosses that involved females from inland Kenya (KI) did not yield any female offspring. However, backcrosses of hybrid female bearing a KI male genome with a KI male yielded both female and male offspring. Thus, there was a partial reproductive incompatibility between KI and West African populations which suggested that the latter were infected with Wolbachia sp. However, this should not affect the efficacy of a population introduced from East into West Africa, as there is a high degree of sib-mating in this gregarious parasitoid species. It was concluded that the regional differences in the relative importance of C. sesamiae was due to differences in the insect and plant host range of the different populations.     

An empirical test of the F2 screen for detection of Bacillus thuringiensis-resistance alleles in tobacco budworm (Lepidoptera: Noctuidae)

Insects exposed to genetically modified crops expressing Bacillus thuringiensis (Bt) toxins are under intense selection pressure that could result on widespread Bt resistance. Screening for early indications of Bt resistance developing in targeted Lepidoptera is conducted in many of the regions where genetically modified cotton and corn have been commercialized. Heliothis virescens (F.) (Lepidoptera: Noctuidae) has been selected in the laboratory to have a gene for resistance to Cry1Ac. We used this laboratory line to test the assumptions and theoretical predictions related to detection of recessive Bt-resistant alleles in field populations based on a second generation (F2) screen. By creating single-pair families from mating a heterozygous Cry1Ac-resistant moth with a Cry1Ac-susceptible moth, we simulated the most common genotype when Bt-resistance alleles are at low frequency in the field. The second generation (F2) neonates of single-pair families were screened daily with diagnostic concentration bioassays. Cry1Ac-resistant homozygous larvae were detected, but the proportion of resistant larvae was generally below the theoretical expectation of 6.25% and was influenced by the moth F1 sib-mating density and by the day of oviposition of F2 eggs. Logistical considerations such as F1 sib-mating density and F2 neonate screening are important for the successful implementation of a reliable method.     

Modification of hormonal balance in larvae of the corn borer Sesamia nonagrioides (Lepidoptera: Noctuidae) due to sublethal Bacillus thuringiensis protein ingestion

Bacillus thuringiensis (Bt) corn, Zea mays L., is highly efficient against the corn borer Sesamia nonagrioides (Lefèbvre) (Lepidoptera: Noctuidae) when the larvae feed only on the transgenic plants. However, when they feed on Bt leaves during only part of their development, thus ingesting sublethal amounts of Bt toxins, some larvae survive. A previous study reported a prolonged development and precocious diapause induction in larvae fed on a diet with sublethal amounts of Cry1Ab protein. To determine whether these effects were accompanied by a modification of the hormonal balance, S. nonagrioides larvae were fed on sublethal amounts of Bt protein provided in Bt leaves or in the diet. The larvae that survived had higher levels of juvenile hormone (JH), whereas their level of ecdysteroids did not increase sufficiently to allow pupation, leading to a longer larval development and more larval molts. This response may be considered a defense mechanism that allows some larvae to survive toxin ingestion; it is similar the response to insecticidal toxins or viruses observed in other larvae. Changes in the hormone levels in diapausing larvae were undetectable, probably because these changes were masked by the higher level of JH in the hemolymph of diapausing larvae and because of lack of ecdysteroid titer increase, a phenomenon that is usually observed a few days before pupation in nondiapausing larvae. These results should be taken into account in the establishment of non-Bt refuges to prevent development of Bt-resistance in S. non-agrioides populations.     

A Stilbene Optical Brightener can Enhance Bacterial Pathogenicity to Gypsy Moth (Lepidoptera: Lymantriidae) and Colorado Potato Beetle (Coleoptera: Chrysomelidae)

Stilbene optical brighteners were first investigated to protect biological control agents such as viruses, fungi, and nematodes against ultraviolet light. Some are known to enhance the activity of insect viruses in Lepidoptera. In this work, one stilbene brightener, Tinopal LPW, also increased mortality of gypsy moth and Colorado potato beetle larvae when treated with bacteria/optical brightener combinations. This increase in mortality, however, did not occur for every bacteria/insect combination. In gypsy moth, a significant increase in larval mortality was observed only with Bacillus thuringiensis combined with Tinopal LPW. In Colorado potato beetle, however, the addition of Tinopal LPW increased larval mortality with all bacteria tested (B. thuringiensis, Serratia marcescens, Photorhabdus luminescens, and Chromobacterium sp.). The brightener also decreased the time to kill for these pathogens. This decrease in LT₅₀ was observed not only for bacteria+Tinopal LPW combinations, but also for combinations of Chromobacterium sp. toxin+Tinopal LPW. The mechanism for increase in bacterial toxicity by optical brighteners is compatible with mechanisms proposed for enhancement based on viral/lepidopteran/optical brightener systems that are not dependent on replication.     

Frequency and severity of western bean cutworm (Lepidoptera: Noctuidae) ear damage in transgenic corn hybrids expressing different Bacillus thuringiensis cry toxins

The frequency and severity of corn ear damage caused by western bean cutworm, Striacosta albicosta (Smith), were measured on transgenic corn, Zea mays L., hybrids expressing two different insecticidal Bacillus thuringiensis (Bt) (Berliner) Cry toxins (Bt) selected to protect against damage caused by larval European corn borer, Ostrinia nubilalis (Hübner). A field cage experiment deliberately infested with western bean cutworm egg masses resulted in less damage in the hybrid expressing the Cry1F protein and supported fewer western bean cutworm larvae than its non-Bt isoline. Corn hybrids expressing Cry1F, grown in small plot field experiments at three locations over two separate years and exposed to natural western bean cutworm infestations suffered less damage than non-Bt or Bt-hybrids expressing a Cry1Ab protein. Later maturing hybrids suffered more damage than shorter-season hybrids. Finally, corn ears observed in strip trials for several years in diverse agronomic conditions in farmer-cooperator fields corroborated the in-plant protection conferred by corn hybrids expressing the Cry1F protein in small plot field trials.     

Inheritance of resistance to Bacillus thuringiensis Cry1Ac toxin in a greenhouse-derived strain of cabbage looper (Lepidoptera: Noctuidae)

A population of cabbage looper, Trichoplusia ni (Hübner), collected from commercial greenhouses in the lower mainland of British Columbia, Canada, in 2001 showed a resistance level of 24-fold to Dipel, a product of Bacillus thuringiensis (Bt) subspecies kurstaki. This population was selected with Cry1Ac, the major Bt Cry toxin in Dipel, to obtain a homogenous population resistant to Cry1Ac. The resulting strain of T. ni, named GLEN-Cry1Ac, was highly resistant to Cry1Ac with a resistance ratio of approximately 1000-fold. The larvae from the GLEN-Cry1Ac strain could survive on Cry1Ac-expressing transgenic broccoli plants that were highly insecticidal to T. ni and diamondback moth, Plutella xylostella (L.). The inheritance of Cry1Ac resistance in this T. ni strain was autosomal and incompletely recessive. The degree of dominance of the resistance was -0.402 and -0.395, respectively, for the neonates in reciprocal crosses between the GLEN-Cry1Ac and a laboratory strain of T. ni. Using chi2 goodness-of-fit test, we demonstrated that the inhibition of larval growth resulting from testing 12 toxin doses in the progeny of the backcross fit the predicted larval responses based on a monogenic inheritance model. Therefore, we conclude that the inheritance of the resistance to Cry1Ac in the T. ni larvae is monogenic.     

Cross-resistance responses of CrylAc-selected Heliothis virescens (Lepidoptera: Noctuidae) to the Bacillus thuringiensis protein vip3A

One susceptible and three Cry1Ac-resistant strains of tobacco budworm, Heliothis virescens (F.) (Lepidoptera: Noctuidae), were used in laboratory studies to determine the level of cross-resistance between the Bacillus thuringiensis (Berliner) toxins Cry1Ac and Vip3A by using concentration-mortality and leaf tissue experiments. Concentration-mortality data demonstrated that the three Cry1Ac-resistant H. virescens strains, YHD2, KCBhyb, and CxC, were at least 215- to 316-fold resistant to Cry1Ac compared with the susceptible strain, YDK. Results from Vip3A concentration-mortality tests indicated that mortality was similar among all four H. virescens strains. Relative larval growth on Cry1Ac reflected concentration-mortality test results, because YHD2 larval growth was mostly unaffected by the Cry1Ac concentrations tested. Growth ratios for KCBhyb and CXC indicated that they had a more moderate level of resistance to Cry1Ac than did YHD2. Relative larval growth on Vip3A was highly variable at lower concentrations, but it was more consistent on concentrations of Vip3A above 25 microg/ml. Differences in larval growth among strains on Vip3A were not as pronounced as seen in Cry1Ac experiments. Mortality and larval growth also was assessed in leaf tissue bioassays in which YDK, CxC, and KCBhyb neonates were placed onto leaf disks from non-Bt and Bt cotton, Gossypium hirsutum L., for 5 d. Three Bt lines were used in an initial bioassay and consisted of two Vip3A-containing lines, COT203 and COT102, and a Cry1Ac-producing line. Mortality of KCBhyb and CXC was lower than that of YDK larvae in the presence of leaf tissue from the Cry1Ac-producing line. Additionally, increased larval growth and leaf tissue consumption on Cry1Ac-containing leaf disks was observed for KCBhyb and CXC. Mortality and larval weights were similar among strains when larvae were fed leaf tissue of either non-Bt, COT203, or COT102. A subsequent leaf tissue bioassay was conducted that evaluated four cotton lines: non-Bt, Cry1Ab-expressing, Vip3A-expressing, and pyramided-toxin plants that produced both Cry1Ab and Vip3A. Mortality levels were similar among strains when fed non-Bt, Vip3A-expressing, or pyramided-toxin leaf tissues. Mortality was higher for YDK than for KCBhyb or CXC on Cry1Ab-expressing leaf tissues. No differences in larval weights were observed among strains for any genotype tested. Results of these experiments demonstrate that cross-resistance is nonexistent between CrylAc and Vip3A in H. virescens. Thus, the introduction of Vip3A-producing lines could delay Cry1Ac-resistance evolution in H. virescens, if these lines gain a significant share of the market.     

Susceptibility of legume pod borer (LPB), Maruca vitrata to delta-endotoxins of Bacillus thuringiensis (Bt) in Taiwan

Baseline susceptibility of legume pod borer (LPB) to the insecticidal crystal proteins (ICPs) from Bacillus thuringiensis, viz, Cry1Aa, Cry1Ab, Cry1Ac, Cry1Ca and Cry2Aa was assessed in Taiwan. Insect bioassays were performed by incorporating the Bt delta-endotoxins into the LPB artificial diet. The efficacy of different Bt delta-endotoxins against second instar larvae of LPB showed that the toxin Cry1Ab was the most potent toxin (LC(50) 0.207ppm), followed by Cry1Ca, Cry1Aa, Cry2Aa and Cry1Ac in descending order, with LC(50)s 0.477ppm, 0.812ppm, 1.058ppm and 1.666ppm, respectively. Hence, Cry1Ab and/or Cry1Ca toxins would provide effective control of early larval stages of LPB.     

Cross-resistance studies of Cry1Ac-resistant strains of Helicoverpa armigera (Lepidoptera: Noctuidae) to Cry2Ab

Bioassays and binding tests between Cry toxins (CrylAa, CrylAb, CrylAc, and Cry2Ab) and brush-border membrane vesicles (BBMVs) from larvae of a Bacillus thurningiensis (Bt)-susceptible (96S) and two CrylAc-resistant strains (BtR and LFR10) were conducted for investigating cross-resistance of CrylAc-resistant strains to Cry2Ab in Helicoverpa armigera (Hiibner) (Lepidoptera: Noctuidae). The resistance ratio (RR) values of the BtR and LFR10 strains to CrylAc and Cry2Ab were 2,971- and 1.1-fold and 253- and 1.0-fold, respectively, indicating that there was no cross-resistance to Cry2Ab. The binding experiments between Cry toxins and BBMVs from BtR, LFR10, and 96S larvae showed that all of the toxins could bind with these BBMVs, but the Cry2Ab could not displace 125I labeled CrylAc and CrylAb. The same results were observed in reciprocal binding tests, demonstrating that CrylA and Cry2Ab had different binding sites in H. armigera and providing a potential mechanism for the lack of cross-resistance between CrylA and Cry2Ab toxins. These results suggest that the transgenic cotton, Gossypium hirsutum L., expressing CrylAc and Cry2Ab genes may be deployed for management of CrylAc resistant H. armigera.     

Differential effects of sugar-mimic alkaloids in mulberry latex on sugar metabolism and disaccharidases of Eri and domesticated silkworms: enzymatic adaptation of Bombyx mori to mulberry defense

Mulberry leaves (Morus spp.) exude latex rich in sugar-mimic alkaloids, 1,4-dideoxy-1,4-imino-d-arabinitol (d-AB1) and 1-deoxynojirimycin (DNJ), as a defense against herbivorous insects. Sugar-mimic alkaloids are inhibitors of sugar-metabolizing enzymes, and are toxic to the Eri silkworm, Samia ricini, a generalist herbivore, but not at all to the domesticated silkworm, Bombyx mori, a mulberry specialist. To address the phenomena, we fed both larvae diets containing different sugar sources (sucrose, glucose or none) with or without sugar-mimic alkaloids from mulberry latex. In S. ricini, addition of sugar-mimic alkaloids to the sucrose (the major sugar in mulberry leaves) diet reduced both growth and the absorption ratio of sugar, but it reduced neither in B. mori. The midgut soluble sucrase activity of S. ricini was low and inhibited by very low concentrations of sugar-mimic alkaloids (IC₅₀=0.9–8.2 μM), but that of B. mori was high and not inhibited even by very high concentrations (IC₅₀>1000 μM) of sugar-mimic alkaloids. In S. ricini, the addition of sugar-mimic alkaloids to the glucose diet still had considerable negative effects on growth, although it did not reduce the absorption ratio of glucose. The hemolymph of S. ricini fed sugar-mimic alkaloids contained sugar-mimic alkaloids. The trehalose concentration in the hemolymph increased significantly in S. ricini fed sugar-mimic alkaloids, but not in B. mori. The trehalase activities of S. ricini were lower and inhibited by lower concentrations of sugar-mimic alkaloids than those of B. mori. These results suggest that sugar-mimic alkaloids in mulberry latex exert toxicity to S. ricini larvae first by inhibiting midgut sucrase and digestion of sucrose, and secondly, after being absorbed into hemolymph, by inhibiting trehalase and utilization of trehalose, the major blood sugar. Further, our results reveal that B. mori larvae evolved enzymatic adaptation to mulberry defense by developing sucrase and trehalase that are insensitive to sugar-mimic alkaloids.     

Effects of MON810 Bt field corn on adult emergence of Helicoverpa zea (Lepidoptera: Noctuidae)

A 3-yr study (1996-1998) was conducted to evaluate the effects of MON810 Bt corn on Helicoverpa zea (Boddie) emergence and to determine whether delayed larval development as a result of Bt intoxication results in higher levels of diapause induction and pupal mortality. In the 1997 study, there was no difference in prepupal mortality between corn types, although significantly more prepupae from Bt plots than from non-Bt plots died in emergence buckets before constructing pupal chambers in 1998. In all years, significantly fewer moths emerged from prepupae collected from Bt plots, suggesting that effects of the expressed Cry1Ab extended to the prepupal and pupal stages. Late plantings of corn showed the greatest reductions in moth emergence from Bt corn because environmental conditions were more conducive to trigger diapause at the time H. zea was developing in these plantings. This was supported by a significantly greater proportion of diapausing pupae remaining in the ground in the late plantings of both Bt and non-Bt corn. For April and early May plantings, larval feeding on Bt corn delayed the time to pupation, although there was no significant difference in moth emergence between corn types for those larvae that successfully pupated. Although Bt expression had less impact on the proportion of moths emerging, the actual number of moths emerging from Bt corn was significantly reduced because fewer larvae reached pupation. Delays in adult emergence, along with significant reductions in adult emergence from MON810 Bt corn, should reduce the rates of colonization in soybean and other late host crops but may also result in asynchrony of mating between individuals emerging from Bt and non-Bt corn. This, in turn, may contribute to the evolution of resistance to Bt corn.     

Binding and toxicity of Bacillus thuringiensis protein Cry1C to susceptible and resistant diamondback moth (Lepidoptera: Plutellidae)

We studied mechanisms of resistance to Bacillus thuringiensis insecticidal crystal protein Cry1C in the diamondback moth, Plutella xylostella (L.). Binding assays with midgut brush border membrane vesicles prepared from whole larvae showed no significant difference between resistant and susceptible strains in binding of radioactively-labeled Cry1C. These results indicate that reduced binding of Cry1C to midgut membrane target sites did not cause resistance to Cry1C. Thus, the mechanism of resistance to Cry1C differs from that observed in several previously reported cases of resistance to Cry1A toxins in diamondback moth. We tested Cry1C toxin and Cry1C crystalline protoxin against resistant and susceptible larvae using leaf disk bioassays. After adjusting for the size difference between Cry1C toxin and protoxin, we found that with resistant larvae, toxin was significantly more toxic than protoxin. In contrast, with susceptible larvae, no significant difference in toxicity occurred between Cry1C toxin and protoxin. The resistance ratios for Cry1C were 19 for toxin and 48 for protoxin. These results suggest that reduced conversion of Cry1C protoxin to toxin is a minor mechanism of resistance to Cry1C. Because neither reduced binding nor reduced conversion of protoxin to toxin appear to be major mechanisms, one or more other mechanisms are important in diamondback moth resistance to Cry1C.     

Sublethal effects of Cry 1F Bt corn and clothianidin on black cutworm (Lepidoptera: Noctuidae) larval development

Black cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae), is an occasional pest of maize (corn), Zea mays L., that may cause severe stand losses and injury to corn seedlings. The efficacy of the neonicotinoid seed treatment clothianidin at two commercially available rates and their interaction with a transgenic corn hybrid (Bt corn), trait expressing the Bacillus thuringiensis variety aizawai insecticidal toxin Cry 1Fa2, against black cutworm larvae was investigated. Clothianidin at a rate of 25 mg kernel(-1) on Bt corn increased larval mortality and reduced larval weight gains additively. In contrast, weights of larvae fed non-Bt corn seedlings treated with clothianidin at a rate of 25 mg kernel(-1) increased significantly, suggesting either compensatory overconsumption, hormesis, or hormoligosis. Both Bt corn alone and clothianidin at a rate of 125 mg kernel(-1) applied to non-Bt corn seedlings caused increased mortality and reduced larval weight gains. In two field trials, plots planted with Bt corn hybrids consistently had the highest plant populations and yields, regardless of whether they were treated with clothianidin at the lower commercial rate of 25 mg kernel(-1) The use of Bt corn alone or in combination with the low rate of clothianidin (25 mg kernel(-1)) seems suitable as a means of suppressing black cutworm in no-tillage cornfields, although rescue treatments may still be necessary under severe infestations. Clothianidin alone at the low rate of 25 mg kernel(-1) is not recommended for black cutworm control until further studies of its effects on larval physiology and field performance have been completed.     

Use of Bacillus thuringiensis Toxins for Control of the Cotton Pest Earias insulana (Boisd.) (Lepidoptera: Noctuidae)

Thirteen of the most common lepidopteran-specific Cry proteins of Bacillus thuringiensis have been tested for their efficacy against newly hatched larvae of two populations of the spiny bollworm, Earias insulana. At a concentration of 100 μg of toxin per milliliter of artificial diet, six Cry toxins (Cry1Ca, Cry1Ea, Cry1Fa, Cry1Ja, Cry2Aa, and Cry2Ab) were not toxic at all. Cry1Aa, Cry1Ja, and Cry2Aa did not cause mortality but caused significant inhibition of growth. The other Cry toxins (Cry1Ab, Cry1Ac, Cry1Ba, Cry1Da, Cry1Ia, and Cry9Ca) were toxic to E. insulana larvae. The 50% lethal concentration values of these toxins ranged from 0.39 to 21.13 μg/ml (for Cry9Ca and Cry1Ia, respectively) for an E. insulana laboratory colony originating from Egypt and from 0.20 to 4.25 μg/ml (for Cry9Ca and Cry1Da, respectively) for a laboratory colony originating from Spain. The relative potencies of the toxins in the population from Egypt were highest for Cry9Ca and Cry1Ab, and they were both significantly more toxic than Cry1Ac and Cry1Ba, followed by Cry1Da and finally Cry1Ia. In the population from Spain, Cry9Ca was the most toxic, followed in decreasing order by Cry1Ac and Cry1Ba, and the least toxic was Cry1Da. Binding experiments were performed to test whether the toxic Cry proteins shared binding sites in this insect. ¹²⁵I-labeled Cry1Ac and Cry1Ab and biotinylated Cry1Ba, Cry1Ia, and Cry9Ca showed specific binding to the brush border membrane vesicles from E. insulana. Competition binding experiments among these toxins showed that only Cry1Ab and Cry1Ac competed for the same binding sites, indicating a high possibility that this insect may develop cross-resistance to Cry1Ab upon exposure to Cry1Ac transgenic cotton but not to the other toxins tested.     

Geographic variation in susceptibility of Chilo suppressalis (Lepidoptera: Pyralidae) to Bacillus thuringiensis toxins in China

Geographic variation in the susceptibility of the striped stem borer, Chilo suppressalis (Walker) (Lepidoptera: Pyralidae), in China to Bacillus thuringiensis (Bt) insecticidal crystal proteins Cry1Ac and Cry1Ab was studied to establish baseline information for comparing the future response of populations with increased exposure to Bt products. Rice is the major host of C. suppressalis, and Bt rice ma) be released in China in the near future. Twelve populations of the pest were collected from the major rice-growing regions of China. LC50 estimates were determined for all populations for Cry1Ac and for eight populations for Cry1Ab. The bioassay results indicated that the range of LC50 in neonate larvae to Cry1Ac and Cry1Ab was from approximately 15 to approximately 157 mg (AI)/L and approximately 2 to approximately 34 mg (AI)/L, respectively. LC50 values were lower for Cry1Ab than for Cry1Ac, and there was a significant positive correlation between the two toxins tested.     

Effectiveness of Bacillus thuringiensis-transgenic chickpeas and the entomopathogenic fungus Metarhizium anisopliae in controlling Helicoverpa armigera (Lepidoptera: Noctuidae)

The use of genetically modified (Bt) crops expressing lepidopteran-specific Cry proteins derived from the soil bacterium Bacillus thuringiensis is an effective method to control the polyphagous pest Helicoverpa armigera. As H. armigera potentially develops resistance to Cry proteins, Bt crops should be regarded as one tool in integrated pest management. Therefore, they should be compatible with biological control. Bioassays were conducted to understand the interactions between a Cry2Aa-expressing chickpea line, either a susceptible or a Cry2A-resistant H. armigera strain, and the entomopathogenic fungus Metarhizium anisopliae. In a first concentration-response assay, Cry2A-resistant larvae were more tolerant of M. anisopliae than susceptible larvae, while in a second bioassay, the fungus caused similar mortalities in the two strains fed control chickpea leaves. Thus, resistance to Cry2A did not cause any fitness costs that became visible as increased susceptibility to the fungus. On Bt chickpea leaves, susceptible H. armigera larvae were more sensitive to M. anisopliae than on control leaves. It appeared that sublethal damage induced by the B. thuringiensis toxin enhanced the effectiveness of M. anisopliae. For Cry2A-resistant larvae, the mortalities caused by the fungus were similar when they were fed either food source. To examine which strain would be more likely to be exposed to the fungus, their movements on control and Bt chickpea plants were compared. Movement did not appear to differ among larvae on Bt or conventional chickpeas, as indicated by the number of leaflets damaged per leaf. The findings suggest that Bt chickpeas and M. anisopliae are compatible to control H. armigera.     

Inheritance of resistance to Bacillus thuringiensis Cry1Ab protein in the sugarcane borer (Lepidoptera: Crambidae)

Inheritance traits of a Cry1Ab-resistant strain of the sugarcane borer, Diatraea saccharalis (F.) were analyzed using various genetic crosses. Reciprocal parental crosses between Cry1Ab-susceptible and Cry1Ab-resistant populations, F₁ by F₁ crosses, and backcrosses of F₁ with the Cry1Ab-resistant population were successfully completed. Larval mortality of the parental and cross-populations were assayed on Cry1Ab diet and Bacillus thuringiensis (Bt)-corn leaf tissue. Maternal effects and sex linkage were examined by comparing the larval mortality between the two F₁ populations. Dominance levels of resistance were measured by comparing the larval mortality of the Cry1Ab-resistant, -susceptible, and -heterozygous populations. Number of genes associated with the resistance was evaluated by fitting the observed mortality of F₂ and backcross populations with a Mendelian monogenic inheritance model. Cry1Ab resistance in D. saccharalis was likely inherited as a single or a few tightly linked autosomal genes. The resistance was incompletely recessive on Bt corn leaf tissue, while the effective dominance levels (D_ML) of resistance increased as Cry1Ab concentrations decreased with Cry1Ab-treated diet. D_ML estimated based on larval mortality on intact Bt corn plants reported in a previous study ranged from 0.08 to 0.26. This variability in D_ML levels of Cry1Ab resistance in D. saccharalis suggests that Bt corn hybrids must express a sufficient dose of Bt proteins to make the resistance genes functionally recessive. Thus, Bt resistant heterozygous individuals can be killed as desired in the “high/dose refuge” resistance management strategy for Bt corn.     

Control of Resistant Pink Bollworm (Pectinophora gossypiella) by Transgenic Cotton That Produces Bacillus thuringiensis Toxin Cry2Ab

Crops genetically engineered to produce Bacillus thuringiensis toxins for insect control can reduce use of conventional insecticides, but insect resistance could limit the success of this technology. The first generation of transgenic cotton with B. thuringiensis produces a single toxin, Cry1Ac, that is highly effective against susceptible larvae of pink bollworm (Pectinophora gossypiella), a major cotton pest. To counter potential problems with resistance, second-generation transgenic cotton that produces B. thuringiensis toxin Cry2Ab alone or in combination with Cry1Ac has been developed. In greenhouse bioassays, a pink bollworm strain selected in the laboratory for resistance to Cry1Ac survived equally well on transgenic cotton with Cry1Ac and on cotton without Cry1Ac. In contrast, Cry1Ac-resistant pink bollworm had little or no survival on second-generation transgenic cotton with Cry2Ab alone or with Cry1Ac plus Cry2Ab. Artificial diet bioassays showed that resistance to Cry1Ac did not confer strong cross-resistance to Cry2Aa. Strains with >90% larval survival on diet with 10 μg of Cry1Ac per ml showed 0% survival on diet with 3.2 or 10 μg of Cry2Aa per ml. However, the average survival of larvae fed a diet with 1 μg of Cry2Aa per ml was higher for Cry1Ac-resistant strains (2 to 10%) than for susceptible strains (0%). If plants with Cry1Ac plus Cry2Ab are deployed while genes that confer resistance to each of these toxins are rare, and if the inheritance of resistance to both toxins is recessive, the efficacy of transgenic cotton might be greatly extended.     

Biology of Campoletis chlorideae (Uchida) (Hym., Ichneumonidae) developing in Bt-treated, Bt-resistant Helicoverpa armigera (Hübner) (Lep., Noctuidae) larvae

Abstract:  Life history parameters in two generations of endoparasitoid Campoletis chlorideae (Uchida) were examined using Bacillus thuringiensis (Bt)-resistant Helicoverpa armigera (Hübner) larvae feeding on B. thuringiensis toxin Cry1Ac. In the laboratory, Bt toxin was fed to Bt-resistant host larvae continuously in case of Bt treatment and only before or after the host larvae were parasitized in Bt–P and P–Bt treatments, respectively. C. chlorideae pupae developed faster in Bt treatment than non-Bt treatment. The shortened pupal stage duration was mainly because of the feeding of host larvae on Bt-diet before being parasitized. Body length of adult male C. chlorideae developed inside Bt-treated Bt-resistant (Bt–Bt) H. armigera larvae significantly decreased, especially in host larvae feeding on Bt-diet after being parasitized. However, survival, pupal mortality and adult longevity of C. chlorideae were almost unaffected in Bt-resistant H. armigera larvae feeding on Bt-toxin. Furthermore, Bt-treated host larvae had the same effect on the F₁ progeny of C. chlorideae as the previous generation, and there was no significant difference between generations. This experiment suggests that there is very limited effect on the life history parameters in two generations of C. chlorideae parasitizing Bt–Bt H. armigera larvae. But both generations of C. chlorideae are affected when Bt-resistant H. armigera larvae fed on Bt toxin for different durations.