Comparative virulence of Beauveria bassiana isolates against lepidopteran pests of vegetable crops

Forty-three isolates of the entomopathogenic fungus Beauveria bassiana were screened for virulence against second-instar larvae of diamondback moth (Plutella xylostella) (DBM), European corn borer (Ostrinia nubilalis) (ECB), corn earworm (Helicoverpa zea) (CEW), and fall armyworm (Spodoptera frugiperda) (FAW); 30 of these isolates were tested against beet armyworm (Spodoptera exigua) (BAW). Highly virulent isolates were also tested against black cutworm (Agrotis ipsilon) (BCW), and the most virulent isolate was also assayed against imported cabbage worm (Pieris rapae) (ICW) and cabbage looper (Trichoplusia ni) (CL). All lepidopteran species tested were susceptible to B. bassiana. Corn earworm and beet armyworm were most susceptible to fungal infection, and fall armyworm was least susceptible. Limited testing suggested low susceptibility of black cutworm and cabbage looper. B. bassiana isolate 1200 exhibited virulence against all pest species greater than or equal to commercial strain GHA of B. bassiana currently registered in the USA as BotaniGard®. In assays in which larvae were topically sprayed and maintained on the treated substrate for 24 h at 100% relative humidity, 6-day (25 °C) median lethal concentrations (LC₅₀s) of this isolate against CEW, BAW, DBM, FAW, ICW, ECB, CL, and BCW were 4, 5, 7, 11, 12, 98, 125, and 273 conidia/mm², respectively. The respective LC₅₀s of commercial strain GHA against these pest species were 9, 67, 97, 1213, 29, 1668, 541, and 3504 conidia/mm². Use of LC₅₀ versus median lethal concentration ratios (comparing LC₅₀s of each isolate to a “standard” strain) generated similar rankings of isolate virulence. Results from parametric ANOVAs of log LC₅₀ values followed by Tukey HSD multiple comparisons tests and those from Kruskal-Wallis nonparametric analyses followed by sequential Bonferroni tests for means comparisons were nearly identical.     

Induction of detoxification enzymes in phytophagous insects: Role of insecticide synergists,larval age,and species

Five insecticide synergists, all of which were either methylenedioxyphenyl compounds or analogs, were compared as to their effect on cytochrome P450 monooxygenase induction caused by an allelochemical in fall armyworm larvae. Feeding the synergists (piperonyl butoxide, safrole, isosafrole, MGK 264, and myristicin) individually to the larvae caused decreases in the microsomal aldrin epoxidase activities ranging from 38% to 74% when compared with controls. Feeding indole-3-carbinol resulted in a 4-fold increase in the microsomal epoxidase activity. However, cotreatment of any of the synergists and the inducer completely eliminated the induction. Sixth instar larvae were more inducible than second instar larvae with respect to microsomal epoxidase and glutathione transferase in the fall armyworm. Enzyme inducibility varied widely among the seven phytophagous Lepidoptera examined. When indole-3-carbinol was used as an inducer of microsomal epoxidase, the extent of inducibility of the enzyme was fall armyworm > velvetbean caterpillar > corn earworm > beet armyworm > tobacco budworm > cabbage looper > diamondback moth. When indole-3-acetonitrile was used as an inducer, the inducibility of glutathione transferase was fall armyworm > beet armyworm > corn earworm > cabbage looper > velvetbean caterpillar > tobacco budworm > diamondback moth. Inducibility of five microsomal oxidase systems also varied considerably in the corn earworm, indicating the multiplicity of cytochrome P450 in this species. Microsomal epoxidase and glutathione transferase were induced by cruciferous host plants such as cabbage and their allelochemicals in diamondback moth larve. © 1993 Wiley-Liss, Inc.     

Conventional resistance of experimental maize lines to corn earworm (Lepidoptera: Noctuidae), fall armyworm (Lepidoptera: Noctuidae), southwestern corn borer (Lepidoptera: Crambidae), and sugarcane borer (Lepidoptera: Crambidae)

Plant resistance is a useful component of integrated pest management for several insects that are economically damaging to maize, Zea mays L. In this study, 15 experimental lines of maize derived from a backcross breeding program were evaluated for resistance to corn earworm, Helicoverpa zea (Boddie); fall armyworm, Spodoptera frugiperda (J. E. Smith); southwestern corn borer, Diatraea grandiosella Dyar; and sugarcane borer, Diatraea saccharalis (F.). Experimental line 100-R-3 was resistant in the field to leaf feeding by fall armyworm and line 116-B-10 was resistant in the field to leaf feeding by fall armyworm and leaf and stalk feeding by southwestern corn borer. When corn earworm larvae were fed field harvested silks from experimental line 81-9-B in the laboratory, their pupal weights were significantly lower than the pupal weights of larvae that were fed silks from the resistant control, Zapalote Chico. Maysin levels lower than those commonly associated with corn earworm resistance were present in the resistant experimental line, 107-8-7, indicating a new basis confers resistance to corn earworm in this line. These resistant experimental lines will provide plant breeders with new sources of resistance to lepidopterous insects for the development of improved maize breeding populations.     

β-Glucosidase in four phytophagous Lepidoptera

β-Glucosidase in larvae of the fall armyworm, Spodoptera frugiperda (J. E. Smith), was studied using helicin as the substrate. Enzyme activity was found in various tissues, with the midgut exhibiting the highest activity. Among the midgut subcellular fractions, the soluble fraction was the most active. The apparent K_m value for the enzyme was 0.63 mM. The enzyme was fairly stable for 8 weeks when stored at −10°C. Activity was mainly located in the 60% fraction when the midgut soluble fraction was fractionated with ammonium sulfate. The system also hydrolyzed a variety of glucosides including numerous toxic plant allelochemicals. Substrate specificity of β-glucosidase was different in the fall armyworm, velvetbean caterpillar, cabbage looper and corn earworm, suggesting a qualitative difference in the enzyme among these species. In general, d-amygdalin, helicin, d(+)-cellobiose, p-nitrophenyl β-d-glucoside and 4-methylumbelliferyl β-d-glucoside were the preferred substrates, whereas sinigrin, phloridzin, α-solanine, tomatine and linamarin were poor substrates for the enzyme in these insects. Toxicity tests reveal that, in most instances, the glucosides were less toxic than their corresponding aglycones to the fall armyworm, indicating that β-glucosidase is an activation enzyme.     

Characterization of induced resistance in tomato plants

We have characterized, using several types of bioassays, the resistance induced in young tomato plants by feeding of the corn earworm, Helicoverpa zea. Beet armyworm larvae, Spodoptera exigua, and leafminers, Liriomyza trifolii, were used to assay the induced resistance. In whole-plant experiments, damage localized to a single leaflet of fourleaf tomato plants induced a systemic increase in resistance such that beet armyworm larvae confined to previously damaged (induced) plants grew at a rate about half that of larvae raised on control plants and consumed less leaf tissue from induced plants than from control plants. In experiments using excised leaves, beet armyworm larvae suffered increased mortality when reared on leaves from induced plants. The strength of this induced resistance varied spatially relative to the damaged position; moreover, the spatial distribution of induced resistance changed over a three-week period following damage. Other experiments demonstrated that the mechanisms of induced resistance in tomato foliage involves both a decrease in larval preference for and a decrease in the nutritional value of induced foliage. Induction also retarded the oviposition and/or early development of leafminers. Thus, induced resistance has relatively severe effects on the biology of subsequent herbivores. These data should allow us to begin to elucidate cause-effect relationships between induced resistance and induced chemistry in tomato plants.     

Establishment and characterization of insect cell lines from 10 lepidopteran species

Summary Cell lines from selected lepidopteran species were established for the overall purpose of use in baculovirus production. A total of 36 new cell lines from 10 lepidopteran species were generated, including cell lines from a pyralid, the European corn borer,Ostrinia nubilalis, a plutellid, the diamondback moth,Plutella xylostella, as well as eight noctuids: the black cutworm,Agrotis ipsilon, the celery looper,Anagrapha falcifera, the velvetbean caterpillar,Anticarsia gemmatalis, the corn earworm,Helicoverpa zea, the tobacco budworm,Heliothis virescens, the beet armyworm,Spodoptera exigua, the fall armyworm,Spodoptera frugiperda, and the cabbage looper,Trichoplusia ni. Tissues used for cell line establishment included fat bodies, ovaries, testes, or whole embryos/larvae/pupae. All the cell lines were subcultured numerous times, characterized by isoenzyme analysis and/or deoxyribonucleic acid amplification fingerprinting using polymerase chain reaction, and stored in liquid nitrogen. Many of the cell lines were adapted to grow in serum-free medium, with cell lines fromA. ipsilon andH. virescens being adapted to suspension culture, using shaker flasks. The potential use for these cell lines in baculovirus production is discussed. All programs and services of the U.S. Department of Agriculture are offered on a nondiscriminatory basis without regard to race, color, national origin, religion sex, age, marital status, or handicap.     

Pathology of a nuclear polyhedrosis virus of the alfalfa looper in alternate hosts

The symptomatology and histopathology of a nuclear polyhedrosis virus isolated from a larva of the alfalfa looper, Autographa californica, was studied by examining 13 tissues in the original and following alternate hosts: cabbage looper, Trichoplusia ni; beet armyworm, Spodoptera exigua; saltmarsh caterpillar, Estigmene acrea; corn carworm, Heliothis zea; cotton leafperforator, Bucculatrix thurberiella; and diamondback moth, Plutella xylostella. In all hosts, the hypodermal, tracheal matrix, and fat body cells were infected. Other tissues infected in some hosts included the Malpighian tubules, muscle, hemocytes, ganglia, midgut, hindgut, juvenile tissue (imaginal buds), and testes. No major changes in tissue tropisms were observed. The external symptoms were typical of nuclear polyhedrosis in all species except the corn earworm; in this host, development of the disease and death were delayed.     

Assessment of experimental Bt events against fall armyworm and corn earworm in field corn

Performance of experimental Bacillus thuringiensis (Bt) MON events alone and pyramided with MON810 were evaluated over 3 yr in Georgia and Alabama. Ability of events to prevent whorl defoliation by the fall armyworm, Spodoptera frugiperda (J. E. Smith), and natural ear feeding damage by the corn earworm, Helicoverpa zea (Boddie) was assessed. In each year, near-isogenic hybrids with novel single transformation events and crosses pyramided with the MON810 event were compared with the standard single MON810 event and nontransformed susceptible control. Events were tested for resistance to whorl damage by manual infestations of fall armyworm and ear damage by natural infestations of corn earworm. All Bt events tested reduced fall armyworm whorl damage ratings per plant compared with the susceptible hybrid. All Bt treatments also had considerably less ear infestation and damage by corn earworm compared with the nontransgenic isoline. The MON841, MION849, and MON851 events reduced ear damage by H. zea but were not as effective as other novel events and were not advanced for further testing after the 1999 season. Pyramiding events compared with single events did not improve control of fall armyworm whorl damage, but they generally did prevent more ear damage by corn earworm. The MON84006 event singly and pyramided with MON810 had superior control of whorl-stage damage by S. frugiperda and ear damage by H. zea compared with MON810. Deployment of new events and genes could provide additional tools for managing the potential for insect resistance to Bt toxins. Furthermore, improved control of whorl and ear infestations by H. zea and S. frugiperda would increase the flexibility of planting corn, Zea mays L., and permit double cropping of corn in areas where these pests perennially reach damaging levels.     

Evaluation of the Spatial Pattern of Steinernema riobravis in Corn Plots

The vertical and horizontal spatial patterns of a naturally occurring population of the entomopathogenic nematode Steinernema riobravis (Rhabditida: Steinernematidae) were investigated in corn field soil by laboratory and field bioassays. This nematode appears to be endemic to the Lower Rio Grande Valley of Texas, where it was found parasitizing prepupae and pupae of both corn earworm, Helicoverpa zea, and fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae). Corn earworm prepupa was the bioassay host used to detect S. riobravis from soil in previously harvested corn plots. Steinernema riobravis occurred at soil depths of 5-30 cm. The maximum nematode density was in the upper 20 cm of soil, and the lowest density occurred at soil depth of 25-30 cm. The field and laboratory bioassays performed on the top 20 era of soil resulted in S. riobravis-infected corn earworm of 49 and 34%, respectively, whereas at 25-30 cm soil depths 11 and 4.5% of the H. zea were infected, respectively. The horizontal spatial pattern of this nematode was patchy or aggregated. Our study provides new information on the spatial pattern of S. riobravis in its natural habitat and indicates the need to augment its natural biocontrol efficacy.     

Toxicity of the fungal metabolite griseofulvin toHelicoverpa zea andSpodoptera frugiperda

Griseofulvin is a fungal metabolite that can be considered a mycotoxin but is used pharmaceutically to treat topical fungal infections. It was found to be toxic to both the corn earworm.Helicoverpa zea Boddie, and the fall armyworm,Spodoptera frugiperda (J. E. Smith), at 250 ppm in diets. Its toxicity was enhanced by exposure to ultraviolet light, suggesting it can be photoactivated. Its toxicity was also enhanced by coincorporated δ-amino levulinic acid, suggesting it interferes with porphyrin synthesis in these insects. Differences in toxicity and rates of metabolism caused by coincorporated piperonyl butoxide suggested unspecific monooxygenases were relatively more important in griseofulvin activation inH. zea and detoxification inS. frugiperda.     

Polymerase chain reaction primers for polymorphic microsatellite loci in the African armyworm,Spodoptera exempta (Lepidoptera: Noctuidae)

Eight pairs of polymerase chain reaction (PCR) primers that amplify polymorphic microsatellite loci were developed for the African armyworm, Spodoptera exempta (Walker) to be used in the study of its population dynamics in sub‐Sahara Africa where the species is a major pest of cereals and rangeland. A magnetic beads based enrichment protocol was used; it appears that previously reported scarcity of microsatellites in Lepidoptera species does not apply to the African armyworm. All the loci showed significant heterozygote deficit; possibly because the samples were laboratory reared from limited stock. Four primer pairs successfully amplified single fragments of beet and fall armyworm DNA of comparable size to the African armyworm alleles.     

Genome mapping coupled with CRISPR gene editing reveals a P450 gene confers avermectin resistance in the beet armyworm

The evolution of insecticide resistance represents a global constraint to agricultural production. Because of the extreme genetic diversity found in insects and the large numbers of genes involved in insecticide detoxification, better tools are needed to quickly identify and validate the involvement of putative resistance genes for improved monitoring, management, and countering of field-evolved insecticide resistance. The avermectins, emamectin benzoate (EB) and abamectin are relatively new pesticides with reduced environmental risk that target a wide number of insect pests, including the beet armyworm, Spodoptera exigua, an important global pest of many crops. Unfortunately, field resistance to avermectins recently evolved in the beet armyworm, threatening the sustainable use of this class of insecticides. Here, we report a high-quality chromosome-level assembly of the beet armyworm genome and use bulked segregant analysis (BSA) to identify the locus of avermectin resistance, which mapped on 15–16 Mbp of chromosome 17. Knockout of the CYP9A186 gene that maps within this region by CRISPR/Cas9 gene editing fully restored EB susceptibility, implicating this gene in avermectin resistance. Heterologous expression and in vitro functional assays further confirm that a natural substitution (F116V) found in the substrate recognition site 1 (SRS1) of the CYP9A186 protein results in enhanced metabolism of EB and abamectin. Hence, the combined approach of coupling gene editing with BSA allows for the rapid identification of metabolic resistance genes responsible for insecticide resistance, which is critical for effective monitoring and adaptive management of insecticide resistance.     

Toxic components on the larval surface of the corn earworm (Heliothis zea) and their effects on germination and growth of Beauveria bassiana

Caprylic acid is present on the surface of corn earworm, Heliothis zea, and fall armyworm, Spodoptera fragiterda, larvae. Because caprylic acid inhibits germination of Beauveria bassiana, presence of this compound will be determined to the establishment of an infection of larvae by this fungus. Other free fatty acids present on the surface of the H. zea and S. frugiterda are tentatively identified as valeric and nonanoic acids; these also possess mycostatic activity toward B. bassiana. Depending on concentration, caprylic acid inhibits germination of conidia for different amounts of time (R. J. Smith and E. A. Grula, 1981, J. Invertebr. Pathol., 37, 222–230). We now further report that inhibition and/or growth is also related to the source of carbon, nitrogen, and energy present in the growth medium. This observation of selective toxicity in the presence of different nutrients was also observed using nonanoic acid. Our data therefore make it necessary to interpret the effects of certain fatty acids on germination and growth of B. bassiana (and probably other fungi as well) in terms of nutrients for the germination process.     

Indirect Chilling Injury of Spodoptera exigua in Response to Long-Term Exposure to Sublethal Low Temperature

We report indirect chilling injury, which is caused by the long-term exposure to the low temperature above freezing point, found in the beet armyworm, Spodoptera exigua. Short (minute or hour) exposure to 5°C was not lethal to all stages of S. exigua. However, several day-exposure to 5°C gave significant nonfreezing injuries such as low egg hatchability, retarded larval development, and low pupation rates. Even though the larvae did not develop at 5°C, they used up their major nutrient reserves significantly. The mechanism of the indirect chilling injury of the beet armyworm was discussed on the basis of the oxidative stress hypothesis.     

甜菜夜蛾不同龄期幼虫对药剂的敏感性及其与酶活力的关系

【目的】明确甜菜夜蛾Spodoptera exigua(Hübner)不同龄期幼虫对甲氨基阿维菌素苯甲酸盐和毒死蜱的敏感性及其与羧酸酯酶、多功能氧化酶、谷胱甘肽S-转移酶和乙酰胆碱酯酶活性的相关性。【方法】采用室内生物测定方法检测甜菜夜蛾不同龄期幼虫对药剂的敏感性,并检测了不同龄期幼虫体内羧酸酯酶、多功能氧化酶、谷胱甘肽S-转移酶和乙酰胆碱酯酶的活力。【结果】在所测定的5个龄期中,随龄期增加,敏感性逐渐降低,其中1~5龄幼虫对甲氨基阿维菌素苯甲酸盐的LC50分别为0.1010、0.3561、0.7568、1.4325和8.4390 mg/L,对毒死蜱的LC50分别为27.4632、46.8495、87.5222、129.3217和1 356.6770 mg/L。单头幼虫的羧酸酯酶、多功能氧化酶、谷胱甘肽S-转移酶和乙酰胆碱酯酶活力随虫龄的增加而提高,与龄期间对药剂的敏感性呈负相关(由于外源化合物摄入量减少,5龄单头幼虫的多功能氧化酶活力略有降低)。【结论】甜菜夜蛾不同龄期幼虫对药剂的敏感性存在非常明显的差异,在田间防治中,应选择对杀虫剂敏感性较高的低龄幼虫作为最佳防治时期。     

甜菜夜蛾群体繁殖及其对杀虫剂毒力敏感性测定

用黄豆粉、麦胚粉、酵母粉为主要营养成份研制成人工饲料,实现了甜菜夜蛾大量饲养及群体繁殖,与自然饲料相比,人工饲料饲养甜菜夜蛾幼虫存活与成虫产卵量明显增加,室内用浸渍法测定11种杀虫剂对甜菜夜蛾2龄幼虫的毒力,结果表明,田间甜菜夜蛾种群对除尽,米满,菜喜最敏感,其次是对抑太保,卡死克,毒丝本,高效灭百可,快灵,对锐劲特,氯氰菊酯,乙酰甲胺磷的毒力均已高度不敏感。     

Noctuid migration in Texas within the nocturnal aeroecological boundary layer

Long-distance migration of adult corn earworm moths (Helicoverpazea), and several other noctuid moth species, facilitates seasonalexpansion of pest populations and consequent increased infestationsof agricultural crops on a continental scale in North America.Long-term field studies of population dynamics and migratoryflights of H. zea and fall armyworm (Spodoptera frugiperda)in the United States were evaluated using X-band radar observationsand profiles of atmospheric conditions. These studies identifiedcharacteristic patterns of migratory flight that are largelyassociated with vertical profiles of temperature and wind speed.Collective patterns of moth migrations were generally highlycorrelated with wind headings, but often at a significant angulardeviation. Preliminary analyses are presented between moth distributionsin the aerosphere estimated from discrete moth counts usingX-band radar and bulk reflectivity data from NEXRAD Dopplerradar. Identification of associations between atmospheric factorsand noctuid population dynamics and migratory flights will improvethe ability to predict infestations by pest species throughouttheir broad seasonal range expansion.     

寄主植物-甜菜夜蛾-寄生蜂三级营养关系的研究进展

综述了近年来甜菜夜蛾Spodoptera exigua (Hübner)、寄主植物和寄生蜂互作关系方面的研究进展。介绍了甜菜夜蛾取食诱导的植物抗虫性产生的原因和机制;阐述了诱导植物产生抗性的甜菜夜蛾激发子volicitin的合成途径和功能,以及虫害诱导的植物挥发物和蛋白酶抑制剂对甜菜夜蛾及其寄生蜂的生态学功能;展望了植物诱导抗虫性在甜菜夜蛾的生物防治和新型抗虫品种开发等领域的应用前景。     

Possible involvement of cyclooxygenase products in the actions of platelet-activating factor and of lipoxygenase products in the vascular effects of epinephrine in perfused rat liver

Platelet-activating factor (PAF) stimulates glycogenolysis and induces vasoconstriction in perfused rat liver. The effect of PAF was rapid but transient and it was blocked by indomethacin and bromophenacyl bromide which suggests a role of cyclooxygenase metabolites in its action. The homologous desensitization of glycogenolysis produced by PAF and the sensitivity of its actions to inhibitors of cyclooxygenase and phospholipase A2 markedly differentiate the mechanism of action of this agent with that of alpha 1-adrenergic agents, vasopressin or angiotensin II. No effect of PAF in isolated hepatocytes was observed which suggest that cells other than hepatocytes could be involved in its action in perfused liver. In addition nordihydroguaiaretic acid and bromophenacyl bromide abolished the vascular effect (but not the glycogenolysis) produced by epinephrine which suggest a role for lipoxygenase products in this effect.