Estigmene Acrea Research Articles

Specific recognition, detoxification and metabolism of pyrrolizidine alkaloids by the polyphagous arctiid Estigmene acrea

Evidence is presented that the polyphagous arctiid Estigmene acrea is well adapted to sequester and specifically handle pyrrolizidine alkaloids of almost all known structural types representative of the major plant families with pyrrolizidine alkaloid-containing species, i.e. Asteraceae with the tribes Senecioneae and Eupatorieae, Boraginaceae, Fabaceae, Apocynaceae and Orchidaceae. The adaptation of E. acrea to pyrrolizidine alkaloids includes a number of specialized characters: (i) highly sensitive recognition of alkaloid sources by pyrrolizidine alkaloid-specific taste receptors; (ii) detoxification of pyrrolizidine alkaloids by N -oxidation catalyzed by a specific flavin-dependent monooxygenase; (iii) transfer and maintenance of all types of pyrrolizidine N-oxides through all developmental stages; (iv) conversion of the various structures into the male courtship pheromone hydroxydanaidal most probably through retronecine and insect specific retronecine esters (creatonotines) as common intermediates; (v) specific integration into mating behavior and defense strategies. Toxic otonecine derivatives, e.g. the senecionine analogue senkirkine, which often accompany the common retronecine derivatives and which cannot be detoxified by N -oxidation do not affect the development of E. acrea larvae. Senkirkine is not sequestered at all. Non-toxic 1,2-saturated platynecine derivatives that frequently occur together with toxic retronecine esters are sequestered and metabolized to hydroxydanaidal, indicating the ability of E. acrea to aromatize saturated pyrrolizidines. Although pyrrolizidine alkaloids, even if they are offered continuously at a high level (2%) in the larval diet, are non-toxic, E. acrea larvae are not able to develop exclusively on a pyrrolizidine alkaloid-containing plant like Crotalaria. Therefore, E. acrea appears to be specifically adapted to exploit pyrrolizidine alkaloid-containing plants as “drug source” but not as a food source.

If you've got it, flaunt it: ingested alkaloids affect corematal display behavior in the salt marsh moth, Estigmene acrea.

Plant-derived pyrrolizidine alkaloids play an important role in the biology of the salt marsh moth, Estigmene acrea (Lepidoptera: Arctiidae). They are phagostimulants for larvae and they stimulate the growth and development of adult male androconial organs called coremata. In this study, we have shown that the pyrrolizidine alkaloid monocrotaline N-oxide (MNO) fed to larvae also affects the courtship behavior of adult males. Males fed a diet containing MNO display their coremata while males fed on the same diet without alkaloid rarely display. This explains why it has been difficult to replicate field observations of the "lekking" behavior of this species in the laboratory where animals are frequently raised on commercially available diets devoid of alkaloids. Corematal inflation was observed in isolated males and in laboratory leks. The effect of larvae feeding on pyrrolizidine alkaloid on the reproductive behavior of adults suggests that this substance may modify the development of the moth's nervous system and contribute to their unusual dual mating strategies. MNO was also shown to be an adequate precursor for the production of the courtship pheromone hydroxydanaidal.

ROLES OF FOOD QUALITY AND ENEMY-FREE SPACE IN HOST USE BY A GENERALIST INSECT HERBIVORE

The relative importance of food quality vs. enemy-free space remains an unresolved but central issue in the evolutionary ecology of host use by phytophagous insects. In this study, we investigate their relative importance in determining host-plant use by a generalist caterpillar, Estigmene acrea (Lepidoptera: Arctiidae). In nature, E. acrea late- instar caterpillars preferred Senecio longilobus (Asteraceae), which contains pyrrolizidine alkaloids that the caterpillars sequester, over Viguiera dentata (Asteraceae), a natal host, and typically suffered a 28% mortality risk from parasitoids. We hypothesized that the natural, mixed diet of caterpillars provides high-quality food via hosts like Viguiera as well as antiparasitoid defense via sequestered toxins from Senecio. We found that a pure Viguiera diet provides superior growth performance over a pure Senecio or mixed diet in the absence of parasitism. However, when parasitism risk is at least moderate, the mixed diet provides a survival advantage over the pure diets of Viguiera or Senecio. We therefore conclude that the balance between benefits of growth (food quality) and defense (enemy-free space) maintains the use of a mixed diet in nature. Furthermore, the value of enemy-free space supercedes the value of food quality in determining the host-plant preference of late-instar caterpillars.

Phenological fate of plant-acquired pyrrolizidine alkaloids in the polyphagous arctiid Estigmene acrea

The alkaloid profiles of the life history stages of the highly polyphagous arctiid Estigmene acrea were established. As larvae individuals had free choice between a plain diet (alkaloid-free) and a diet that was supplemented with Crotalaria-pumila powder with a known content and composition of pyrrolizidine alkaloids (PAs). Idiosyncratic retronecine esters (insect PAs) accounted for approximately half of the PAs recovered from the larvae. These alkaloids were synthesized by the larvae through esterification of dietary supinidine yielding the estigmines, and esterification of retronecine yielding the creatonotines. The retronecine is derived from insect-mediated degradation of the sequestered pumilines (macrocyclic PAs of the monocrotaline type). With one exception, the PA profiles established for larvae were found almost unaltered in all life-stages as well as larval exuviae and pupal cocoons. The exception is the males, which in comparison to pupae and adult females, showed a significantly decreased quantity of the creatonotines and pumilines. These data support the idea that the creatonotines are direct precursors of the PA-derived male courtship pheromone, hydroxydanaidal. Crosses of PA-free males with PA-containing females and vice versa confirmed an efficient trans-mission of PAs from males to females and then from females to eggs. In single cases a male bestowed almost his total PA load to the female, and a female her total load to the eggs. The results are discussed with respect to pheromone formation, PA transmission between life-stages, and the defensive role of PAs against predators and parasitoids

Foraging in nature: foraging efficiency and attentiveness in caterpillars with different diet breadths

Abstract. 1. Seventy‐seven individual last‐instar caterpillars foraging in the field were examined for 6 h each. They represented four species of Arctiidae of similar size and habitat use. Two, Hypocrisias minima and Pygarctia roseicapitis, are specialists restricted to particular plant genera. The other two, Grammia geneura and Estigmene acrea, are extreme generalists that use many host plant species from multiple plant families.2. Parameters of behavioural efficiency were monitored. Generalists spent more time walking, rejected more potential host plants, took longer to decide to feed after inspecting a plant, and took relatively more small feeding bouts compared with specialists.3. This is the first test of differential foraging efficiency in the field in relation to diet breadth of insects and the data indicate that generalists are less efficient in their foraging activities and may suffer from divided attention. The need for attentiveness to enhance efficiency and thereby reduce ecological risk is discussed.

Diel periodicity and influence of age and mating on female sex pheromone titre in Estigmene acrea (Lep., Arctiidae)

Abstract: Calling behaviour, diel periodicity, and effect of age and mating on female sex pheromone titre in Estigmene acrea (Drury) were studied under laboratory conditions. Forty‐five per cent of females started calling during the first scotophase, but the highest number of calling females was observed during the second, third and fourth scotophases. Calling behaviour occurred from the third hour after dark until just before the end of the scotophase. However, females exhibited a bimodal pattern of calling with the first peak occurring between 4 and 6 h and a second peak at 10 h after the onset of scotophase. The mean onset of calling time differed significantly with age. Older females showed a tendency to call longer, but there was no significant difference. The amount of (Z,Z)‐3,6‐cis‐9, 10‐epoxyheneicosadiene in females was quantified from the first scotophase following emergence, until the fifth scotophase. Glands of 0‐day‐old females presented a higher content of pheromone compared with that found in glands of 1‐, 2‐, 3‐ and 4‐day‐old females. Pheromone titre was determined at 2‐h intervals throughout the third scotophase and photophase. (Z,Z)‐3,6‐cis‐9,10‐epoxyheneicosadiene was found in the gland during the scotophase as well as the photophase. However, there was no consistent pattern of pheromone production throughout the scotophase or photophase. Mated females of E. acrea produced significantly less pheromone than virgin females.

Acquired and partially de novo synthesized pyrrolizidine alkaloids in two polyphagous arctiids and the alkaloid profiles of their larval food-plants.

The profiles of pyrrolizidine alkaloids (PAs) in the two highly polyphagous arctiids Estigmene acrea and Grammia geneura and their potential PA sources in southeastern Arizona were compiled. One of four species of Boraginaceae, Plagiobothrys arizonicus, contained PAs; this is the first PA record for this plant species. The principle PA sources are Senecio longilobus (Asteraceae) and Crotalaria pumila (Fabaceae). The known PA pattern of S. longilobus was extended; the species was found to contain six closely related PAs of the senecionine type. Three novel PAs of the monocrotaline type, named pumilines A-C, were isolated and characterized from C. pumila, a species not studied before. The pumilines are the major PAs in the seeds, while in the vegetative organs they are accompanied by the simple necine derivatives supinidine and as the dominant compound subulacine (1beta,2beta-epoxytrachelanthamidine). In both plant species, the PAs are stored as N-oxides, except C. pumila seeds, which accumulate the free bases. Great variation in PA composition was observed between local populations of C. pumila. The PA profiles were established for larvae and adults of E. acrea that as larvae had fed on an artificial diet supplemented with crotalaria-powder and of G. geneura fed with S. longilobus. In both experiments, the larvae had a free choice between the respective PA source and diet or food plants free of PAs. The profiles compiled for the two species reflect the alkaloid profiles of their PA sources with one exception, subulacine could never be detected in E. acrea. Besides acquired PAs, insect PAs synthesized from acquired necine bases and necic acids of insect origin were detected in the two arctiid species. These insect PAs that do not occur in the larval food sources accounted for some 40-70% (E. acrea) and 17-37% (G. geneura) of total PAs extracted from the insects. A number of novel insect PAs were identified. Plant-acquired and insect PAs were found to accumulate as N-oxides. The results are discussed in relation to specific biochemical, electrophysiological, and behavioral mechanisms involved in PA sequestration by arctiids.

Dietary alkaloids and the development of androconial organs in Estigmene acrea

Male salt marsh moths, Estigmene acrea (Lepidoptera: Arctiidae), possess inflatable androconial organs called coremata. Prior to mating males form aggregations and inflate their coremata en masse. The communal display attracts additional males and females for the purpose of mating. The coremata are known to carry the plant-derived dihydropyrrolizine, hydroxydanaidal. This pheromonal substance is derived from secondary plant chemicals called pyrrolizidine alkaloids found in the larval diet. When E. acrea larvae were raised on semi-synthetic diets containing different levels of the pyrrolizidine alkaloid precursors the alkaloids triggered a pronounced morphogenetic effect. Adult males that fed on high levels of the pyrrolizidine alkaloid monocrotaline N-oxide (2500 microg) developed the largest coremata. Males that fed on lower levels of monocrotaline N-oxide (500 microg) or no alkaloid, while normal in body weight, had coremata that were progressively smaller and less robust. The size of the coremata and their commensurate pheromonal charge may have behavioral consequences in the unusual mating system of this species.

Dietary alkaloids and the development of androconial organs in Estigmene acrea

Male salt marsh moths, Estigmene acrea (Lepidoptera: Arctiidae), possess inflatable androconial organs called coremata. Prior to mating males form aggregations and inflate their coremata en masse. The communal display attracts additional males and females for the purpose of mating. The coremata are known to carry the plant-derived dihydropyrrolizine, hydroxydanaidal. This pheromonal substance is derived from secondary plant chemicals called pyrrolizidine alkaloids found in the larval diet.When E. acrea larvae were raised on semi-synthetic diets containing different levels of the pyrrolizidine alkaloid precursors the alkaloids triggered a pronounced morphogenetic effect. Adult males that fed on high levels of the pyrrolizidine alkaloid monocrotaline N-oxide (2500 µg) developed the largest coremata. Males that fed on lower levels of monocrotaline N-oxide (500 µg) or no alkaloid, while normal in body weight, had coremata that were progressively smaller and less robust. The size of the coremata and their commensurate pheromonal charge may have behavioral consequences in the unusual mating system of this species.

Are insect-synthesized retronecine esters (creatonotines) the precursors of the male courtship pheromone in the arctiid moth Estigmene acrea?

The pyrrolizidine alkaloid (PA) profiles were determined for adults of the polyphagous arctiid Estigmene acrea, which as larvae had fed on artificial diet supplemented with Crotalaria-pumila powder with known concentrations of PAs. The larvae always had a free choice between alkaloid-containing and plain diets. The alkaloid profiles of adults revealed a striking sexual dimorphism. Both sexes contained macrocyclic PAs of the monocrotaline type sequestered from the diet and, in addition, a substantial proportion of supinidine and retronecine esters synthesized by the insects from necine bases derived from the dietary alkaloids and necic acids of insect origin. These insect alkaloids accounted for 35% and 55% of total PAs in males and females, respectively. The difference was that in females the retronecine esters (creatonotines) made up 58 microg (43% of total PAs), while males contained a fivefold lower proportion, 12 microg (13%). Four of the ten male individuals analyzed were found devoid of creatonotines. Based on the experimental data in combination with evidence from the literature, it is suggested that the creatonotines are direct pheromone precursors in E. acrea. It is hypothesized that this may represent a general mechanism of hydroxydanaidal formation from diverse macrocyclic PAs in arctiids.

A taste receptor neurone dedicated to the perception of pyrrolizidine alkaloids in the medial galeal sensillum of two polyphagous arctiid caterpillars

Abstract.It is shown that pyrrolizidine alkaloids are phagostimulants for the caterpillars of two polyphagous arctiid caterpillars, Estigmene acrea and Grammia geneura. The caterpillars will also eat dry glass‐fibre discs containing only pyrrolizidine alkaloid − an example of pharmacophagy. The tip‐recording technique is used to demonstrate that each species has a neurone in the medial galeal styloconic taste sensillum responding to pyrrolizidine alkaloids, although the species differ in their sensitivities. This neurone responds to at least four different pyrrolizidine alkaloids and their N‐oxides, and experiments indicate that it is dedicated to perception of these compounds. The sensory response is phasic–tonic and during the tonic phase remains unchanged for at least 500 ms, resembling the type of response often seen in neurones that are sensitive to plant secondary compounds producing deterrent effects.

Field efficacy and seasonal expression profiles for terminal leaves of single and double Bacillus thuringiensis toxin cotton genotypes.

Examination of commercial Cry1Ac transgenic Bacillus thuringiensis Berliner (Bt) cotton varieties (Bollgard, Monsanto, St. Louis, MO) and an experimental Cry1Ac + Cry2Ab transgenic Bt cotton variety (Bollgard II, Monsanto) for lepidopteran field efficacy was conducted during the 2000 growing season. In addition, a commercially available (Envirologix, Portland, ME) quantification assay (ELISA) was used to measure and profile the expression levels of Cry proteins in two of these varieties ['DP50B, Bollgard'; 'DP50BII, Bollgard II' (Delta & Pine Land, Scott, MS)]. Populations of beet army worms, Spodoptera exigua (Hübner), and soybean loopers, Pseudoplusia includens (Walker), were significantly lower (P < 0.05) in Bollgard II plots compared with Bollgard. Population numbers for fall army worms, Spodoptera frugiperda (J. E. Smith), and salt marsh caterpillars, Estigmene acrea (Drury), were lower in Bollgard II plots compared with Bollgard but means did not differ significantly (P > 0.05). Single and dual-toxin genotypes remained superior (P < 0.05) compared with conventional cotton against the tobacco budworm, Heliothis virescens (F.). The addition of Cry2Ab had no significant (P > 0.05) impact on Cry1Ac expression in Bollgard II compared with Cry1Ac expression in Bollgard. Furthermore, throughout the season Cry2Ab was present at much higher levels in the plant compared with Cry1Ac for Bollgard II plants. Possible species-specific reasons for increased efficacy of Bollgard II over Bollgard are discussed.

The use of lectins to select subpopulations of insect cells

Lectins have been used in glycoprotein purification, oligosaccharide analysis, and in cell-selection processes. Here, we utilize lectins in a rational attempt to select a subpopulation of insect cells (Estigmene acrea, EAA) with more complete glycosylation capacity by selecting cells that display more complex-type cell-surface oligosaccharides than the general population of cells. A lectin (ECA) from Erythrina cristagalli, specific for galactose beta(1-4)N-acetylglucosamine, was found to be useful in recognizing a small subpopulation of Sf-21 and EAA cells. Cell selections were performed by lectin affinity chromatography and by selective agglutination. Analysis by lectin blots of cell lysates and a quantitative agglutination assay did not reveal significant differences in regard to the level of complex glycosylation between the negatively and positively selected subpopulations of EAA cells. Statistically significant differences in binding the fluorescently labeled lectin, ECA-TRITC were observed even 30 passages post-selection between EAA subpopulations that were negatively and positively selected by lectin affinity chromatography. There were no differences in the two subpopulations in the ECA quantitative agglutination assay. Thus, the hypothesis that a subpopulation differing in glycosylation capacity exists and that such a subpopulation can be identified by the character of cell-surface oligosaccharides is plausible. However, these differences appear to be too small to be of practical use. Copyright 1999 John Wiley & Sons, Inc.

Growth of Paenibacillus larvae, the causative agent of American foulbrood in honey bees and Bacillus popilliae, the causative agent of milky disease in beetle larvae in lepidopteran cell cultures

AbstractTNM‐FH Lepidopteran insect cell culture medium containing 10% fetal bovine serum (FBS), while allowing limited vegetative growth of Paenibacillus larvae (wild‐type strain), the causative agent of American foulbrood, contained no viable vegetative cells upon subculture, nor were any heat resistant spores produced in this medium alone. However, TNM‐FH medium cotaining embryonic or midgut cells from Trichoplusia ni, hemocytes from Estigmene acrea, ovarian and embryonic cells from Spodoptera frugiperda, embryonic cells from Plutella xylostella, Spodoptera exigua and Pseudaletia unipuncta or ovarian cells from Lymantria dispar, supported both heavy vegetative cell growth and moderate production of heat resistant spores. EX‐CELL 405 serum‐free insect cell culture medium alone appeared to contain the appropriate nutrients required for both vegetative growth and sporulation of P. larvae. However, in the presence of embryonic cells from T. ni, limited vegetative growth occurred and the P. larvae cells appeared to die off. This was confirmed by the fact that no colony growth occurred upon subculture, nor were any heat resistant spores detected. This was true also in the presence of fat body cells from T. ni, except that a limited number of spores (4,000/ml) were detected in the form of cology‐forming units (CFU) on plates following heating to 80°C for 20 minutes. In a parallel study with a wild‐type strain of Bacillus popilliae, vegetative cells grew only in TNM‐FH medium in the presence of mid‐gut BTI‐Tn‐MG and ovarian (Tn‐368) cells of T. ni. No heat resistant spores, however, were detected in any of the cultures. When BTI‐Tn‐MG and Tn‐368 cells were further challenged with four variant cultures of B. popilliae, vegetative growth and limited sporulation were achieved. The BTI‐Tn‐MG cell line in TNM‐FH medium produced as many as 12,000 spores/ml after 21 days in culture.

N-acetyl-beta-glucosaminidase accounts for differences in glycosylation of influenza virus hemagglutinin expressed in insect cells from a baculovirus vector.

The hemagglutinin of fowl plague virus has been expressed in Spodoptera frugiperda (Sf9) cells and in Estigmene acrea cells by using a baculovirus vector. Structural analysis revealed that the endo-H-resistant N-glycans of HA from Sf9 cells were predominantly trimannosyl core oligosaccharides, whereas in E. acrea cells most of these cores were elongated by at least one terminal N-acetylglucosamine residue. To understand the difference in carbohydrate structures, enzymes involved in N-glycan processing have been analyzed. The results revealed that the different glycosylation patterns observed are due to an N-acetyl-beta-glucosaminidase activity that was found in Sf9 cells but not in E. acrea cells. This enzyme specifically used the GlcNAcMan(3)GlcNAc(2) oligosaccharide as a substrate. When N-acetyl-beta-glucosaminidase or alpha-mannosidase II was inhibited by specific inhibitors, the amount of terminal N-acetylglucosamine in hemagglutinin from Sf9 cells was significantly enhanced. These results demonstrate that N glycosylation in both cell lines follows the classical pathway up to the stage of GlcNAcMan(3)GlcNAc(2) oligosaccharide side chains. Whereas these structures are the end product in E. acrea cells, they are degraded in Sf9 cells to Man(3)GlcNAc(2) cores by N-acetyl-beta-glucosaminidase.

Biochemical Analysis of the N-Glycosylation Pathway in Baculovirus-Infected Lepidopteran Insect Cells

The baculovirus—insect cell system is used routinely for foreign glycoprotein production, but the precise nature of the N-glycosylation pathway in this system remains unclear. Some studies indicate that these cells cannot process N-linked oligosaccharides to complex forms containing outer-chain galactose and sialic acid, while others indicate that they can. In this study, we used the major virion envelope glycoprotein of the baculovirus Autograpba californica multicapsid nuclear polyhedrosis virus (AcMNPV) to probe the N-glycosylation pathway in baculovirus-infected lepidopteran insect cells. The results showed that gp64 contained mannose, fucose, and probably N -acetylglucosamine, but no detectable galactose or sialic acid. These same results were observed with gp64 produced in any one of three different lepidopteran insect cell lines derived from Spodoptera frugiperda, Trichoplusia ni , or Estigmene acrea, whether it was produced at relatively earlier or later times after infection. These results indicated that the gp64 produced in AcMNPV-infected lepidopteran insect cells lacks complex N-linked oligosaccharides containing outer-chain galactose and sialic acid. By contrast, gp64 produced in mammalian cells contained both galaclose and sialic acid, and endoglycosidase digestions revealed that these sugars were constituents of N-linked, not O-linked, oligosaccharides. This showed that at least one N-linked side chain on gp64 has the potential to be processed to a complex form. Together, these results suggest either that AcMNPV-infected lepidopteran insect cells are unable to convert any of the N-linked side chains on gp64 to complex structures or that outer-chain galactose and sialic acid residues are added to gp64 and then removed by cellular or viral exoglycosidases.

Replication and morphogenesis of Amsacta moorei entomopoxvirus in cultured cells of Estigmene acrea (salt marsh caterpillar)

A study of the sequence of morphogenic events occurring within Amsacta moorei entomopoxvirus-infected Estigmene acrea cells is presented. Stages in virion development, and the various cytopathic effects observed in these cells between 0 and 120 h post-infection (p.i.) are described. Events in the early stages of virion assembly (24 to 48 h p.i.), the formation of the outer viral membrane (48 to 72 h p.i.) and the development of occlusion bodies or spheroids (72 to 120 h p.i.) were identified. Cells grown in TC100 culture medium supported production of mature virus particles, the majority of which were either of the intracellular naked virion form, or mature virions incorporated into occlusion bodies. Only limited production of the extracellular enveloped form was observed in these cells.

Comparative recombinant protein production of eight insect cell lines

A recombinant Autographa californica baculovirus expressing secreted alkaline phosphatase (SEAP) gene was used to evaluate the expression of a secreted glycoprotein in eight insect cell lines derived from Spodoptera frugiperda, Trichoplusia ni, Mamestra brassicae and Estigmene acrea. Because cell density was found to influence protein production, SEAP production was evaluated at optimal cell densities for each cell line on both a per cell and per milliliter basis. On a per cell basis, the T. ni-derived BTI-TN-5B1-4 cells produced a minimum of 20-fold more SEAP than the S. frugiperda-derived Sf9 or Sf2l cell lines and a minimum of 9-fold more than any of the other cell lines growing in serum-containing medium. On a per milliliter basis, BTI-TN-5B1-4 cells produced a minimum of fivefold more SEAP than any of the other cell lines tested. Using cell lines that were adapted to serum-free medium, SEAP yields were the same or better than their counterparts in serum-containing medium. At 3 days postinoculation, extracellular SEAP activity ranged from 59 to 85% of total SEAP activity with cell lines grown in serum-free and serum-containing media.

Cytopathic effects of Amsacta moorei entomopoxvirus infection on the cytoskeleton of Estigmene acrea cells

Virus-induced rounding of cultured Estigmene acrea cells infected with Amsacta moorei entomopoxvirus (AmEPV), observed by phase contrast and scanning electron microscopy, indicated that virus infection resulted in reorganization of the host cell cytoskeleton. Immunofluorescent staining of the tubulin and f-actin components showed a progressive contraction of both the microtubule and microfilament network during the period of AmEPV infection. Microtubules began to depolymerize between 12 and 24 h post-infection (h p.i.) and by 48–72 h p.i., the tubulin had further contracted to form a reduced network around the main cell body, whilst the f-actin became rearranged to form distinct foci and microspikes. During the later stages of infection (96 h p.i.), localization of both tubulin and f-actin could be detected in the area of cytoplasm associated with virus assembly. By 120 h p.i., virus spheroids were detected in the cytoplasm and the tubulin and f-actin were reduced to sparse patches over the cell surface. Depolymerization of the microtubules by colchicine corresponded to the virus-mediated effects, and virus replication was not impeded by colchicine-induced depolymerization. Once virus-induced polymerization of f-actin had occurred to form foci, these remained throughout the period of infection. Treatment of virus-infected cells with aphidicolin and cycloheximide indicated that these effects may be mediated by both early and late virus genes.

Resistance of Cotton Lines Containing a Bacillus thuringiensis Toxin to Pink Bollworm (Lepidoptera: Gelechiidae) and other Insects

Three transgenic lines of cotton, Gossypium hirsutum L., carrying a modified insect-control protein from Bacillus thuringiensis subsp. kurstaki (Berliner), were evaluated for resistance to severallepidopterous insects. These three lines, along with the explant source cultivar, ‘Coker 312’, and a locally adapted control, ‘MD 51 ne’, were grown in a field experiment at Maricopa, Arizona. Early in the season, before bolls were available for infestation, the number of rosetted blooms caused by pink bollworm, Pectinophora gossypiella (Saunders), was 95% lower in the transgenic lines than in the control cultivars. The pink bollworm larvae penetrated bolls of the transgenic lines readily; however, live larvae recovered from incubated bolls, and percent seed damage were reduced 97-99% in the transgenic lines compared with the control cultivars. No live larvae were recovered from bolls of two transgenic lines, 62 and 65. The transgenic lines were highly resistant to cotton leaf perforator, Bucculatrix thurberiella Busck, as shown by an absence of larval populations and of leaf mining and feeding. The transgenic lines sustained very little leaf damage from saltmarsh caterpillar, Estigmene acrea (Drury), and beet armyworm, Spodoptera exigua (Hubner), whereas the control cultivars were almost defoliated by season’s end. As expected, the transgenic lines were not resistant to several nonlepidopterous insect pests, and they did not affect populations of beneficial insects. Higher populations of sweetpotato whitefly, Bemisia tabaci (Gennadius), on the transgenic lines than on the control cultivars may have been a consequence of reduced leaf feeding damage (by lepidopterous insects) rather than increased whitefly susceptibility of the transgenic lines. A breeding strategy to increase the insect resistance of cotton plants would be to combine the bacterial toxin trait with other resistance traits such as nectariless, okra leaf, and early maturity, known to reduce pink bollworm and other pest insects.