Characterization and ecological implications of midgut proteolytic activity in larvalPieris rapae andTrichoplusia ni

In their larval luminal midgut fluid,Trichoplusia ni (Lepidoptera: Noctuidae) andPieris rapae (Lepidoptera: Pieridae) contain endopeptidases as their primary proteases. Neither species has detectable exopeptidase activity. Studies using enzyme-specific substrates and inhibitors demonstrate that the endopeptidases are serine proteinases (both trypsinlike and chymotrypsinlike) with histidine at the active site. Optimal pH for the tryptic and chymotryptic activity is 8.5 and 8.0, respectively, forT. ni. and 8.0 and 9.0, respectively, forP. rapae. The efficiency of proteolytic digestion (as measured by the rate of in vitro digestion of a standard protein by the midgut luminal fluid) is positively correlated with the larval dietary protein requirement and is significantly influenced by the ratios of tryptic to chymotryptic activity present in the gut lumen of these two species of Lepidoptera.     

Potent natural egg-laying stimulant for cabbage butterflyPieris rapae

Solutions of glucobrassicin (3-indolylmethyl glucosinolate) purified from foliage and sinigrin (allyl glucosinolate) elicited oviposition by the cabbage butterfly,Pieris rapae, at threshold concentrations as low as 10^–6 M. At higher concentrations, glucobrassicin elicited a faster oviposition rate and a stronger visual response to the substrate through associative learning. Solutions of 10^–5 M glucobrassicin and 10^–2 M sinigrin stimulated equally. Their enzymic hydrolysis products failed to influence oviposition. The markedly greater potency of glucobrassicin is consistent with known glucosinolate profiles of crucifers which show indolyl glucosinolates predominant in foliage.     

Plant secondary compounds as oviposition deterrents for cabbage butterfly,Pieris rapae (Lepidoptera: Pieridae)

Oviposition byPieris rapae butterflies was deterred by spraying the plant secondary compounds coumarin and rutin on cabbage plants in greenhouse choice tests. In no-choice tests ranging from 5 min to 24 hr, acceptance of rutin-treated plants for oviposition increased with trial duration. Both coumarin and rutin deterred oviposition primarily by affecting prealighting rather than postalighting behavior, indicating that deterrence was mediated by noncontact cues.     

Genetic aspects of interpopulational differences in pheromone blend of cabbage looper moth,Trichoplusia ni

The genetic basis of interpopulational differences in the pheromone blend emitted by the cabbage looper moth,Trichoplusia ni (Hübner), was examined by crossing individuals from a field-derived population (P₁) with individuals from a long-maintained laboratory colony (P₂). These colonies differed in the emission rate and relative proportions of four of the five known minor pheromone components, but not in the emission rate of the major component, (Z)-7-dodecenyl acetate (Z7-12Ac). These differences in pheromone blend were quantitatively small but biologically significant, because in the field, males responded preferentially to traps baited with a pheromone blend that is similar to that emitted by P₁ females relative to a blend similar to that emitted by P₂ females. In initial crosses, variation in the quantity and quality of pheromone blends among families of P₁, P₂, and F₁ hybrid females was examined. In F₁ females the relative proportions (quantity relative to the major component) of (Z)-5-dodecenyl acetate (Z5-12Ac) and (Z)-7-tetradecenyl acetate (Z7-14Ac) were intermediate to parental lines. In a second more extensive set of crosses, analyses included P₁, P₂, F₁, F₂, and selected backcrosses. The relative proportion of Z5-12Ac, Z7-14Ac, and Z9-14Ac emitted by F₁ females were intermediate to parental lines. The frequency distributions of relative proportions of these components emitted by females were not consistent with those expected under a single autosomal or sex-linked gene hypothesis, suggesting that more than one gene is involved in the quantitative differences in the pheromone blend.     

Isolation and identification of oviposition deterrents to cabbage butterfly,Pieris rapae,fromErysimum cheiranthoides

Avoidance of some crucifer species by the crucifer specialist,Pieris rapae, has been attributed to the presence of oviposition deterrents in these plants. Studies on one such unacceptable plant,Erysimum cheiranthoides, have resulted in the isolation ofn-butanol-soluble deterrents from the alcoholic extract of foliage. The active fraction contained three cardiac glycosides, which were isolated by reversed-phase HPLC and by open column chromatography on silica gel. Chemical and spectral evidence (UV, [¹H]NMR, and FAB-MS) led to the characterization of these compounds as erysimoside (1), erychroside (2), and erycordin (3). Erysimoside and erychroside were strongly deterrent toPieris rapae, but erycordin was inactive. Both active compounds have the same aglycone, strophanthidin (5) and the inner sugar in both cases is a 2,6-dideoxy hexose to which the outer sugar is attached at position C-4. These structural features, which are absent in the inactive compound (3), may represent specific requirements for oviposition deterrent activity.     

Dodecyl acetate,a second pheromone component of the cabbage looper moth,Trichoplusia ni

Dodecyl acetate was identified as a second component of the sex pheromone ofTrichoplusia ni (Hübner). Dodecyl acetate comprised 9.6% by weight of the total pheromone [(Z)-7-dodecenyl acetate plus dodecyl acetate] extracted from glands and 7.3% by weight of the total pheromone evaporated from the surfaces of glands. Dodecyl acetate appears to function as a short-range pheromone component. Evaporation at female release rates of a 1090 mixture of dodecyl acetate and (Z)-7-dodecenyl acetate in the field caused a significantly greater percentage of males to land on the pheromone source, increased significantly the time they spent on the source, but decreased significantly the time they spent searching for the source when within 50 cm, as compared to (Z)-7-dodecenyl acetate alone.Lepidoptera: Noctuidae.     

Pheromone hydrolysis by cuticular and interior esterases of the antennae,legs, and wings of the cabbage looper moth,Trichoplusia ni (Hübner)

Examination was made of the hydrolytic activities of esterases obtained from the antennae, legs, and wings of 3-day-old cabbage looper moths,Trichoplusia ni (Hübner), by elution and by homogenation of those appendages. Pheromone hydrolysis in 1-min assays was monitored by use of tritium-labeled (Z)-7-dodecen-1-ol acetate and thin-layer chromatography to separate the reaction products. Listed according to the activities of the esterases obtained by homogenation, the organs were antennae > legs > wings. In contrast, the order according to the activities of the eluted esterases was wings > legs > antennae. Also, the eluted enzymes were less active than the esterases obtained by homogenization. The relatively high pheromone-hydrolyzing activity present in homogenized antennae suggests that the esterases originated inside the antennae and lends support to the hypothesis proposed in earlier investigations that pheromone-inactivating enzymes may play an important role in the olfactory process, possibly by clearing pheromone from the vicinity of the olfactory receptors. The esterases detected on the cuticle, on the other hand, may function by preventing surface accumulation of pheromone. The higher measured esterase activity in homogenates of prothoracic legs than of mesothoracic or metathoracic legs suggests that the prothoracic legs, which are used to clean the antennae of debris, may function by removing and degrading pheromone from the surface of antennae.Mention of a commercial or proprietary product does not constitute an endorsement by the USDA.     

Chemical constituents of an unacceptable crucifer,Erysimum cheiranthoides,deter feeding byPieris rapae

The wild cruciferErysimum cheiranthoides was found to contain extractable constituents that deterred feeding by larvae of the crucifer specialistPieris rapae when applied to cabbage leaf disks in both choice and nochoice bioassays. High-performance liquid chromatography was used to separate the extract into several fractions, two of which retained the feeding deterrent activity of the extract. UV-absorption spectra of the fractions suggested that one contained cardenolides similar or identical to those reported to deter oviposition byP. rapae onE. cheiranthoides. The other active fraction evidently contains a compound that deters larval feeding but not adult oviposition. The results suggest that the chemical defense ofE. cheiranthoides depends on two types of compounds acting on separate developmental stages of the insect.     

Compound interactions effects of plant antioxidants in combination with carbaryl on performance ofTrichoplusia ni (Cabbage Looper)

Plant chemicals naturally exist in complex mixtures, which can interact either additively, synergistically, or antagonistically. We investigated the potential interactions of three naturally occurring antioxidants— nordihydroguaiaretic acid (NDGA), safrole, and -tocopherol—with the general insecticide carbarayl to affect the performance of cabbage looper larvae (Trichoplusia ni). The cabbage looper is known to produce a mixed-function oxidase enzyme system in response to the presence of carbaryl. We proposed that plant antioxidants would interfere with enzymatic oxidation, enhancing the susceptibility of this insect to carbaryl. Insects were fed artificial diets containing each antioxidant alone or in pairwise combinations with the insecticide carbaryl to test for their effects on the insect's nutritional measurement indices. The three antioxidants tested were not equally effective individually against insect survivorship and interacted differentially in combination with the insecticide. The nutritional indices were measured on insects fed diets containing the chemicals at nonlethal doses. Insects fed 0.001 % wet wt NDGA diets grew 1.62 times less, and had gross and net conversion efficiencies reduced 3.20 and 3.63 times, respectively, compared to the control larvae. Carbaryl (0.002% wet wt) in combination with NDGA acts as an antagonist to the effects mentioned above, while safrole (1 × 10^–4 wet wt) had an additive effect when combined with the insecticide, reducing 1.76 times larval relative growth rate and efficiency of conversion of ingested food in respect to the control. The larvae fed significantly more (1.2 times) on both insecticide and safrole diets than on the controls or their combined diets. Larvae fed -tocopherol alone or in combination with carbaryl had similar growth and conversion efficiencies as controls. We conclude that the effects of different combinations of compounds cannot be predicted a priori and must be determined experimentally.Operated for the U.S. Department of Energy by the University of California under Contract No. DE-ACO3-76-SF00012. This article was supported by the Center for Intermedia Transport Research (EPA Contract CR-81172-01) and by the Ecological Research Division of the Office of Health and Environmental Research, U.S. Department of Energy.     

Behavioral and physiological responses of cabbage looper,Trichoplusia ni (Hübner), to steam distillates from resistant versus susceptible soybean plants

Soybean plant volatiles, extracted as steam distillates, significantly affected the behavior and biology of the cabbage looper,Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae). Distillates from the susceptible Davis variety attractedT. ni larvae and female adults, whereas those from resistant PI 227687 plants repelled them. When mixed in an artificial diet, steam distillates from PI 227687 plants caused mortality of first-instar larvae. Adults emerging from pupae topically treated with 5 g of such PI 227687 extractable showed developmental abnormalities. Larval feeding was significantly less on Davis leaves treated with PI 227687 volatiles as compared to solvent (acetone) or such Davis extractables. However, Davis volatiles on PI 227687 leaves did not increase larval feeding. HPLC analyses of steam distillates from susceptible Davis versus resistant PI 227687 indicated differences.     

A mutation in pheromonal communication system of cabbage looper moth,Trichoplusia ni

A mutation that results in a dramatic change in the relative proportions of the pheromone components produced by female cabbage looper moths has been found. The most notable changes involve reduction in the emission rate of the major component [(Z)-7-dodecenyl acetate], near absence of a component [(Z)-5-dodecenyl acetate] that is normally present at about 20% of the major component, and a remarkable ca. 20-fold increase in a trace component [(Z)-9-tetradecenyl acetate]. In spite of the multiplicity of changes in the pheromone blend, a genetic analysis indicates that the condition is controlled by a single autosomal gene. These mutants are ineffective in attracting conspecifics, but do attract another distantly related noctuid moth. These results suggest that the evolutionary process that leads to distinct chemical signals in sibling species may include single gene mutations that lead to major changes in the species-specific blend.     

Effects of host plant,Gossypium hirsutum L., on sexual attraction of cabbage looper moths,Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae)

Unmated female or male cabbage looper moths,Trichoplusia ni (Hübner), were attracted more often in a flight tunnel to a cage with moths of the opposite sex and a bouquet of cotton foliage. Increased sexual attractiveness of females with plants may be a result of stimulation of pheromone release in response to plant odor, since more males were attracted when odor of cotton foliage was passed over females than when odor of females was passed over cotton foliage before venting into the flight tunnel. Increased sexual attractiveness of males with plants is due in part to host odor enhancement of female attraction to male pheromone, since more females were attracted to synthetic male pheromone (a blend of enantiomers of linalool and isomers of cresol) and a cotton leaf extract than were attracted to male pheromone alone. A short synthesis procedure was developed for (S)-(+)-linalool, the major component of the male sex pheromone, isolated from hair pencils, used in these tests.     

Comparison of electroantennograms from female and male cabbage looper moths (Trichoplusia ni) of different ages and for various pheromone concentrations

The electroantennogram (EAG) responses of both male and femaleTrichoplusia ni to two components of the female-released sex pheromone were determined over a range of concentrations of chemical and age of insect. The EAG was at a peak at a dose of 100 g for both dodecyl acetate and (Z)-7-dodecenyl acetate (Z7–12OAc) for both sexes. The EAG responses varied with age of the insect. Responsiveness to both components was at a maximum at three days postemergence for males followed by a fairly rapid senescence, and for females it peaked at three days followed by a rapid senescence forZ7–12OAc but not for dodcecyl acetate.     

Plant and fungal cell wall fragments activate expression of proteinase inhibitor genes for plant defense

Plant and fungal cell wall fragments produced by enzymic degradation during pest attacks are hypothesized to be activators of a universal recognition system for locally and systemically activating genes which control the synthesis of plant defense chemicals such as the antibiotic phytoalexins and antinutritive proteinase inhibitors. Proteinase inhibitor cDNAs have been prepared from wound-induced mRNAs, isolated, and characterized. The cDNAs have been utilized to quantify specific proteinase inhibitor mRNAs in leaves following wounding or simulated insect attacks. The cDNAs have also been utilized as hybridization probes to isolate and characterize proteinase inhibitor genes from tomato and potato genomic DNA. Proteinase inhibitor proteins have been induced in tomato leaves by chewing insects and shown to be highly correlated with a systemically mediated reduction in the nutritional quality of the leaves toward the larval noctuidSpodoptera exigua. Thus, the wound-induced proteinase inhibitors, whose genes in tomato leaves can be activated by wounding, insect attacks, and plant and fungal cell wall fragments, can significantly decrease the quality of the leaves for such herbivorous insects. This inducible set of biochemical reactions leading to the de novo biosynthesis of proteinase inhibitors is, therefore, considered to be a potentially important defense of plant leaves that should be considered both in developing general theories on insect-plant interactions and in selecting insect-resistant crop varieties.     

Stratospheric ozone depletion and plantinsect interactions: Effects of UVB radiation on foliage quality ofCitrus jambhiri forTrichoplusia ni

Projected decreases in stratospheric ozone may result in increases in shortwave ultraviolet (UVB) irradiation at the earth's surface. Furanocoumarins, phototoxic compounds found inCitrus jambhiri foliage, increase in concentration when these plants are grown under enhanced UVB. Survivorship schedules ofTrichoplusia ni (Lepidoptera: Noctuidae) caterpillars reared on plants in the presence and absence of enhanced UVB regimes differ significantly; larvae develop more slowly in early life when reared on plants exposed to increased UVB. This same developmental pattern is observed whenT. ni larvae are reared on artificial diets amended with ecologically appropriate amounts of furanocoumarins. Thus, anthropogenically derived changes in stratospheric ozone and concomitant changes in UV light quality at the earth's surface may influence ecological interactions between insects and their host plants by altering secondary metabolism and hence foliage quality for herbivores.     

Influence of trichome exudates from species ofLycopersicon on oviposition behavior ofHeliothis zea (Boddie)

Cage experiments revealed that accessions of the wild tomato speciesLycopersicon hirsutum were preferred sites for oviposition byHeliothis zea. Hexane extracts from the leaves ofL. hirsutum were also preferred sites of oviposition in choice experiments among extracts from severalLycopersicon species. Extracts ofL. hirsutum were still biologically active several days after application, indicating that the phytochemical(s) involved are relatively stable and of low volatility. Gas Chromatographic analysis of leaf hexane extracts from 12 different accessions of theL. hirsutum complex and three tomato cultivars revealed substantial qualitative and quantitative variation in the chemical composition of these extracts. Comparison of these results with extract oviposition studies implicate a group of structurally related compounds as the active agents. Mass spectroscopy has tentatively identified these compounds as sesquiterpenes with the chemical formula C₁₅H₂₂O₂. These compounds are apparently synthesized and secreted from glandular trichomes on the leaf surface. These phytochemicals did not stimulate ovipositional behavior in females of the cabbage looper,Trichoplusia ni. The existence of genetic variation for the presence and amount of kairomones that serve as cues for insect orientation and oviposition could be utilized in a breeding program to develop tomato cultivars with genetically modified allelochemic profiles that would disrupt the sequential behavioral processes of insect host-plant selection.     

Influence of foliar glucosinolates in oilseed rape and mustard on feeding and growth of the bertha armyworm,Mamestra configurata Walker

The relationship between host plant glucosinolate profile and feeding and growth of the Bertha armyworm,Mamestra configurata Walker was investigated using eight cultivated rape and mustard varieties. Mean larval weights of neonates reared on intact rosette-stage plants were significantly different on the different species in the orderBrassica juncea <Sinapis alba <B. napus <B. campestris. WhileB. juncea was least preferred,S. alba was significantly more attractive to neonate larvae in choice tests. Relative consumption and growth rates of fourth-instar larvae were also reduced onB. juncea foliage. Other differences were dependent on the plant growth stage. Neonate preference was not correlated to total glucosinolate levels, but rather to the concentrations of isothiocyanate-releasing glucosinolates. However, the relationship between consumption and glucosinolate levels was inconsistent. Relative growth rate was negatively correlated to total glucosinolate content for stage 3 and 4 foliage—mainly due to the concentration of isothiocyanatereleasing glucosinolates. The relative importance of isothiocyanate-releasing glucosinolates was verified by rearing neonates on meridic diets containing equimolar concentrations of sinigrin, its metabolite, allyl isothiocyanate, and indole-3-carbinol, metabolite of 3-indolylmethyl glucosinolate. Sinigrin and allyl isothiocyanate in the diet produced virtually identical negative weight vs. concentration regression lines. No such dose-response effect was observed with indole-3-carbinol. The data suggest that foliar isothiocyanate-releasing glucosinolates may provide some degree of plant protection from polyphagous insects.