Effects of plant nutrition on the balance of insect relevant cardenolides and glucosinolates in Erysimum cheiranthoides

The possible effects of environmental stress on plant chemistry that are important to herbivorous insects were examined by growing a wild crucifer, Erysimum cheiranthoides, under different nutrient regimes. Oviposition by the cabbage butterfly, Pieris rapae, is thought to be affected by the balance of glucosinolates (stimulants) and cardenolides (deterrents) at the surface of leaves. E. cheiranthoides seedlings were provided with three levels of nitrogen and two levels of sulfur for a period of 15 days before analysis of semiochemicals in whole leaf tissue and at the surface of the foliage. The ratio of cardenolides to glucosinolates in the plants at elevated C/N ratios followed the carbon/nutrient balance hypothesis. However, a high nitrogen supply enhanced biomass production to the extent that concentrations of secondary compounds were unchanged or reduced. The concentration of glucosinolates (glucoiberin and glucocheirolin) at the surface was positively related to whole tissue levels. However, cardenolide (erysimoside and erychroside) concentrations, which were highest in leaf tissue of nitrogen-deficient plants, had the lowest surface levels on foliage of these plants. Possible reasons for differential expression of cardenolides and glucosinolates in a plant as a result of nutrient deficiency are discussed.     

Plant secondary substances and insects behaviour

Allelochemicals are storing in different location in plant tissues as inactive form. Number of identified compounds may now exceed 100,000. Environmental factors have an effect on allelochemicals concentration in plants. Many allelochemicals classified as toxins or deterrents for herbivorous insects. Allelochemicals play a major role in feeding or ovipositing stimulants for some specialist insects. Consumption and assimilation of herbivorous insects had affected by the type of allelochemicals in host plants. Allelochemicals have an acute or chronic toxicity on herbivorous insects. Most specialist herbivorous insects rely heavily of ingested plant allelochemicals. Plant allelochemicals may influence an insect's susceptibility to pathogens such as bacteria, fungi and nematode. Specialists herbivorous insect can be using the allelochemicals in their host plants as protection against natural enemies. Some herbivorous insects are synthesising the aggregation, attracting, alarm or mating pheromone from the allelochemicals in their host plants.     

Differential selection of host plants by twoPieris species: the role of oviposition stimulants and deterrents

Oviposition responses ofPieris rapae L. andP. napi oleracea Harris to nine crucifers, one Capparidaceae and one Tropaeolaceae were directly compared under controlled conditions. Chemical fractions from these plants were also tested on both insects for the presence of oviposition stimulants or deterrents. The results showed that plant chemistry is a key factor in differential selection of potential hosts by thesePieris species. Some plant species were equally acceptable to bothPieris species. However,P. rapae preferred cabbage over most test plants whereasP. napi oleracea strongly preferred plant species that were avoided byP. rapae. The observed preferences were explained in most cases by the presence of stimulants and deterrents in extracts of the plants. The twoPieris species have apparently evolved differential sensitivities to the chemical stimuli that trigger or deter oviposition. The balance of positively and negatively interpreted sensory signals evoked by plant chemicals obviously plays an important role in acceptance or rejection of a plant by both species. The role of specific glucosinolates and differing structure-activity relationships is suggested.     

Effects of nonhost and host plants on insect herbivory covarying with plant size in the cruciferous plant <Emphasis Type="Italic">Turritis glabra</Emphasis>

Correlation between plant size and reproductive output may be modified by herbivory in accordance with host plant density and the presence of nonhost plants. To elucidate the effects of nonhost plant density and host plant density on the intensity of herbivory and reproductive output of the host plant in relation to plant size under natural conditions, we investigated the abundance of three lepidopteran insects, Plutella maculipennis, Anthocharis scolymus, and Pieris rapae the intensity of herbivory, and fruit set of their host plant, Turritis glabra (Cruciferae). To elucidate the effects of nonhost and host plant density, we selected four categories of plots under natural conditions: low density of nonhost and high density of host plants; low density of both nonhost and host plants; high density of both nonhost and host plants; and high density of nonhost and low density of host plants. The plant size indicated by stem diameter was a good predictor of the abundance of all herbivorous species. The effects of density of nonhost and host plants on the abundance of insects varied among species and stages of insects. As the abundance of insects affected the intensity of herbivory, herbivory was more apparent on larger host plants in plots with low density of both nonhost and host plants. Consequently, the correlation between plant size and the number of fruits disappeared in low plots with density of both nonhost and host plants. In this T. glabra– herbivorous insect system, the density of nonhost plants and host plants plays an important role in modifying the relationship between plants and herbivores under natural conditions. Received: July 19, 1999 / Accepted: June 15, 2000     

A trypsin inhibitor from <Emphasis Type="Italic">Cassia obtusifolia</Emphasis> seeds: isolation,characterization and activity against <Emphasis Type="Italic">Pieris rapae</Emphasis>

A trypsin inhibitor was isolated from Cassia obtusifolia by ammonium sulfate precipitation, Sepharose 4B-trypsin affinity and Sephadex G-75 chromatography. The inhibitor consisted of a single polypeptide chain with a molecular mass of 19, 812.55 Da. It was stable from pH 2 to 12 for 24 h, whereas it was unstable either above 70°C for 10 min or under reduced conditions. The inhibitor, which inhibited trypsin activity with an apparent Ki of 0.3 μM, had one reactive site involving a lysine residue. The native inhibitor was resistant to pepsin digestion, whereas the heated inhibitor produced 40% degree of susceptibility. The disulfide linkage and lysine residue were important in maintaining its conformation. Partial amino acid sequence of the purified protein showed a high degree of homology with various members of the Kunitz inhibitor family. Moreover, the inhibitor showed significant inhibitory activity against trypsin-like proteases present in the larval midgut on Pieris rapae and could suppress the growth of larvae.     

Infochemically mediated tritrophic interaction webs on cabbage plants

In response to damage by herbivores, plants are known to emit infochemicals that enhance the effectiveness of insect parasitoids. Studies on plant–parasitoid interactions mediated by such infochemicals have focused on the tritrophic systems in which plants are infested by a single herbivore species. In natural ecosystems, however, plants are often simultaneously infested by several herbivorous species. The present study focuses on two herbivorous species that simultaneously attack crucifer plants and their respective parasitic wasps. We first show the specific responses of the two specialist parasitic wasps [Cotesia plutellae and C. glomerata (Hymenoptera: Braconidae)] to infochemicals originating from cabbage plants (Brassica oleracea cv. Sikidori) infested by each of their respective host larvae [Plutella xylostella (Lepidoptera: Yponomeutidae) and Pieris rapae (Lepidoptera: Pieridae)]. We then coupled the two tritrophic systems on the same cabbage plants. These experiments demonstrated the presence of indirect interactions between the two species of herbivores. Overall, the results indicate the presence of infochemically mediated tritrophic interaction webs on a single plant. Received: September 1, 2000 / Accepted: February 8, 2001     

Kunitz-type trypsin inhibitor with high stability from <Emphasis Type="Italic">Spinacia oleracea</Emphasis> L. seeds

The trypsin inhibitor SOTI was isolated from Spinacia oleracea L. seeds through ammonium sulfate precipitation, Sepharose 4B-trypsin affinity chromatography, and Sephadex G-75 chromatography. This typical Kunitz inhibitor showed remarkable stability to heat, pH, and denaturant. It retained 80% of its activity against trypsin after boiling for 20 min, and more than 90% activity when treated with 6 M guanidine hydrochloride. The formation of stable SOTI-trypsin complex (K _i = 2.3·10⁻⁶ M) is consistent with significant inhibitory activity of SOTI against trypsin-like proteinases present in the larval midgut of Pieris rapae. Sequences of SOTI fragments showed homology with other inhibitors. Published in Russian in Biokhimiya, 2009, Vol. 74, No. 1, pp. 131–140.     

The influence of management practice on the spatial distribution of Lepidopteran pests in <Emphasis Type="Italic">Brassica</Emphasis> crops in the Democratic People’s Republic of Korea: implications for sequential sampling plans

Plutella xylostella and Pieris rapae are the key components of a pest complex that attacks Brassica crops in the Democratic People’s Republic of Korea (DPRK). We examined the spatial distributions of these insects within crops both as individual species and when combined as a standard insect that was derived from their relative feeding rates. The influence of standard co-operative management practice and an integrated pest management (IPM) strategy on the dispersion of the standard insect was tested. Iwao’s m* − m relation was then used to describe the distribution of standard insects by management categories and of Pieris rapae using all data. Pest management practices only affected the distribution of the species when they were combined into standard insects. Enumerative sampling plans were therefore designed for standard insects based on population data derived from IPM-managed fields and for Pieris rapae from population data from all experimental fields. The presented plans have the potential to make a significant contribution to managing lepidopteran pests in the DPRK. The approach will be useful in the design of sequential sampling plans for other geographical regions where these pests co-occur and can also contribute to the development of sequential sampling plans for other pest complexes for which standard insects can be derived.     

A review of the pre-oviposition behaviour of small cabbage white butterfly, Pieris rapae(lepidoptera: Pieridae)

This paper reviews the literature concerning the ovipositional behaviour of Pieris rapae (small cabbage white butterfly) and where appropriate considers results from studies involving other butterflies. The paper considers searching behaviour, stimuli utilised in host plant finding and identification and concludes with a general section on the role of learning and prior experience in ovipositional behaviour.     

The use of oviposition‐induced plant cues by Trichogramma egg parasitoids

1. Female parasitoids have evolved various foraging strategies in order to find suitable hosts. Egg parasitoids have been shown to exploit plant cues induced by the deposition of host eggs. 2. The tiny wasp Trichogramma brassicae uses oviposition‐induced cues from Brussels sprouts to locate eggs of the cabbage white butterflies Pieris brassicae and Pieris rapae that differ in their egg‐laying behaviour. These plant cues are elicited by male‐derived anti‐aphrodisiac pheromones in the accessory reproductive gland (ARG) secretions of mated female butterflies. However, the closely related generalist species Trichogramma evanescens does not respond to Brussels sprout cues induced by the deposition of P. brassicae egg clutches. 3. Here we showed in two‐choice bioassays that T. evanescens wasps respond to Brussels sprout cues induced by (i) the deposition of single eggs by P. rapae, and (ii) the application of ARG secretions from either mated P. rapae females, or from virgin female butterflies in combination with P. rapae's anti‐aphrodisiac compound indole. The wasps only associatively learned to respond to Brussels sprout cues after applying indole alone by linking those cues with the presence of P. rapae eggs. 4. Our results indicate that Trichogramma wasps more commonly exploit oviposition‐induced plant cues to locate their host eggs. Generalist wasps show less specificity in their response than specialists and employ associative learning.     

Midgut cysteine protease-inhibiting activity in Trichoplusia ni protects the peritrophic membrane from degradation by plant cysteine proteases

The action of plant cysteine proteases on the midgut peritrophic membrane (PM) of a polyphagous herbivorous lepidopteran, Trichoplusia ni, was studied. Proteins in PMs isolated from T. ni larvae were confirmed to be highly resistant to the serine proteinases trypsin and chymotrypsin, but were susceptible to degradation by plant cysteine proteases, which is consistent with the known molecular and biochemical characteristics of the T. ni PM proteins. However, the PM proteins were not degraded by plant cysteine proteases in larvae or in the presence of larval midgut fluid in vitro. With further biochemical analysis, cysteine protease-inhibiting activity was identified in the midgut fluid of T. ni larvae. The cysteine protease-inhibiting activity was heat resistant and active in the tested pH range from 6.0 to 10.0, but could be suppressed by thiol reducing reagents or reduced by treatment with catalase. In addition to T. ni, cysteine protease-inhibiting activity was also identified from two other polyphagous Lepidoptera species, Helicoverpa zea and Heliothis virescens. In conclusion, results from this study uncovered that herbivorous insects may counteract the attack of plant cysteine proteases on the PM by inhibiting the potentially insecticidal cysteine proteases from plants in the digestive tract. However, the biochemical identity of the cysteine protease-inhibiting activity in midgut fluid has yet to be identified.     

Solubilization, Activation, and Insecticidal Activity of Bacillus thuringiensis Serovar thompsoni HD542 Crystal Proteins

Cry15Aa protein, produced by Bacillus thuringiensis serovar thompsoni HD542 in a crystal together with a 40-kDa accompanying protein, is one of a small group of nontypical, less well-studied members of the Cry family of insecticidal proteins and may provide an alternative for the more commonly used Cry proteins in insect pest management. In this paper, we describe the characterization of the Cry15Aa and 40-kDa protein's biochemical and insecticidal properties and the mode of action. Both proteins were solubilized above pH 10 in vitro. Incubation of solubilized crystal proteins with trypsin or insect midgut extracts rapidly processed the 40-kDa protein to fragments too small to be detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, whereas the Cry15 protein yielded a stable product of approximately 30 kDa. Protein N-terminal sequencing showed that Cry15 processing occurs exclusively at the C-terminal end. Cry15 protein showed in vitro hemolytic activity, which was greatly enhanced by preincubation with trypsin or insect gut extract. Larvae of the lepidopteran insects Manduca sexta, Cydia pomonella, and Pieris rapae were susceptible to crystals, and presolubilization of the crystals enhanced activity to P. rapae. Activity for all three species was enhanced by preincubation with trypsin. Larvae of Helicoverpa armigera and Spodoptera exigua were relatively insensitive to crystals, and activity against these insects was not enhanced by prior solubilization or trypsin treatment. The 40-kDa crystal protein showed no activity in the insects tested, nor did its addition or coexpression in Escherichia coli increase the activity of Cry15 in insecticidal and hemolytic assays.     

Receptor based models for spontaneous colour choices in flies and butterflies

Colour is one of several stimuli used by herbivorous insects in host choice. Insects have between 2 and 5 different types of photoreceptors to catch quanta of different wavelengths of the spectrum. Many insects have been shown to possess opponent neural interactions between the receptors that enable them to see colour. I present simple models to describe colour choices as functions of the receptor quantum catches and linear interactions of the receptor types. Models are applied to data sets obtained from own experiments and from the literature, on Pieris brassicae and P. rapae (Lepidoptera, Pieridae), Papilio aegeus (Papilionidae), Dacus oleae (Diptera, Tephritidae) and Eristalis tenax (Syrphidae). In fruit flies, detection of green fruit is based on an inhibitory interaction between a green-sensitive receptor type and a blue-sensitive receptor type. This might explain the preference many herbivorous insects have for yellow over green stimuli. Pollen feeding in hoverflies might have evolved from yellow pollen being a super-normal stimulus for herbivorous insects. In butterflies, an additional red-receptor is involved in the colour choice for an oviposition substratum and leads to them choosing green and not yellow. The models introduced in this study open new perspective for a physiological understanding of the design of visual stimuli for monitoring and trapping pest insects.     

Impact of foliar herbivory on the development of a root-feeding insect and its parasitoid

The majority of studies exploring interactions between above- and below-ground biota have been focused on the effects of root-associated organisms on foliar herbivorous insects. This study examined the effects of foliar herbivory by Pieris brassicae L. (Lepidoptera: Pieridae) on the performance of the root herbivore Delia radicum L. (Diptera: Anthomyiidae) and its parasitoid Trybliographa rapae (Westwood) (Hymenoptera: Figitidae), mediated through a shared host plant Brassica nigra L. (Brassicaceae). In the presence of foliar herbivory, the survival of D. radicum and T. rapae decreased significantly by more than 50%. In addition, newly emerged adults of both root herbivores and parasitoids were significantly smaller on plants that had been exposed to foliar herbivory than on control plants. To determine what factor(s) may have accounted for the observed results, we examined the effects of foliar herbivory on root quantity and quality. No significant differences in root biomass were found between plants with and without shoot herbivore damage. Moreover, concentrations of nitrogen in root tissues were also unaffected by shoot damage by P. brassicae larvae. However, higher levels of indole glucosinolates were measured in roots of plants exposed to foliar herbivory, suggesting that the development of the root herbivore and its parasitoid may be, at least partly, negatively affected by increased levels of these allelochemicals in root tissues. Our results show that foliar herbivores can affect the development not only of root-feeding insects but also their natural enemies. We argue that such indirect interactions between above- and below-ground biota may play an important role in the structuring and functioning of communities.     

Oryzacystatin I expressed in transgenic potato induces digestive compensation in an insect natural predator via its herbivorous prey feeding on the plant

We observed recently that the rice cysteine proteinase inhibitor, oryzacystatin I (OCI) expressed in transgenic potato does not affect growth and development of the two-spotted stinkbug predator (Perillus bioculatus) via its herbivorous prey feeding on the plant. Here we monitored the inhibitory activity of recombinant OCI along this potato --> herbivore --> predator continuum, to determine if the absence of effect was associated with a digestive compensatory response of the predator following inhibition of its proteinases by the recombinant cystatin. After confirming that OCI is present in the plant, and ingested in an active form by potato beetle larvae, quantitative and electrophoretic assays allowed us to determine that the recombinant cystatin (representing about 0.8% of total soluble proteins in leaves) was entirely bound to a approximately 30-kDa target proteinase in the prey's midgut, forming a sodium dodecyl sulphate (SDS)-stable complex detected on immunoblots with an anti-OCI polyclonal antibody. Despite the apparent absence of free, residual OCI in the beetle's midgut, digestive protease activity in the predator, known to include OCI-sensitive activity, was altered negatively when the prey was fed the modified plant. This inhibitory process at the third trophic level was accompanied by a compensatory response in the predator, by which serine-type proteinases were synthesized de novo. Overall, our data suggest that the affinity between OCI and the predator's OCI-sensitive proteinases is: (i) as strong as (or stronger than) the affinity between OCI and the potato beetle 30-kDa-sensitive proteinase; and (ii) stronger than the affinity between these enzymes and the plant endogenous homologue of OCI, potato multicystatin, induced in the plant by potato beetle feeding. Our results also show that predatory organisms can adapt their digestive metabolism to the presence of plant antidigestive proteins ingested by their herbivorous preys. In a broader context, this study stresses the need to monitor the inhibitory effects of PI-expressing plants not only on the herbivorous insects targeted, but also on the organisms likely to consume these pests in the environment.     

Constituents of host- and non-host plants deterring oviposition by the cabbage butterfly, Pieris rapae

Oviposition by Pieris rapae L. on cabbage was deterred by homogenized cabbage tissue sprayed onto intact plants. Ether extracts of cabbage also were deterrent, but water extracts were not. Hexane extracts of other host plants deterred oviposition and water extracts had little or no effect. Polar as well as non-polar extracts of non-host plants inhibited oviposition. Polar deterrents in the non-host crucifers, Erysimum cheiranthoides and Capsella bursa-pastoris may explain the avoidance of these plants by P. rapae. Chemical deterrents, as well as stimulants, apparently play a major role in the acceptance or rejection of plants as hosts by ovipositing female butterflies.

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Zusammenfassung Die Eiablage von Pieris rapae auf Kohl wurde gehemmt von einem Homogenisat aus Kohlblättern, das auf intakte Pflanzen gesprüht wurde. Ether-Extrakte der Kohlblätter wirkten im Gegensatz zu Wasserextrakten abschreckend. Ebenso zeigten Hexanextrakte anderer Wirtspflanzen eine hemmende Wirkung auf die Eiablage, während Wasserextrakte anderer Wirtspflanzen keine Wirkung aufwiesen. Dagegen aber verhinderten sowohl polare als auch unpolare Extrakte von Nicht-Wirtspflanzen die Eiablagen. Polare, abschreckende Substanzen in den Nicht-Wirtspflanzen Erysimum cheiranthoides und Capsella bursa-pastoris (Cruciferae) könnten Ursache dafür sein, dass P. rapae diese Pflanzen meidet. Chemische Substanzen mit sowohl abschreckender als auch stimulierender Wirkung spielen offensichtlich eine bedeutende Rolle für legebereite Schmetterlingsweibchen bei der Auswahl von Pflanzen als geeigneten Eiablageplatz.

     

Biochemical characterization of midgut digestive proteases from Mamestra brassicae (cabbage moth; Lepidoptera: Noctuidae) and effect of soybean Kunitz inhibitor (SKTI) in feeding assays

Proteolytic activities in soluble protein extracts from Mamestra brassicae (cabbage moth) larval midgut were analysed using specific peptide substrates and proteinase inhibitors. Serine proteinases were the major activities detected, with chymotrypsin-like and trypsin-like activities being responsible for approximately 62% and 19% of the total proteolytic activity towards a non-specific protein substrate. Only small amounts of elastase-like activities could be detected. The serine proteinases were active across the pH range 7-12.5, with both trypsin-like and chymotrypsin-like activities maximal at pH 11.5. The digestive proteinases were stable to the alkaline environment of the lepidopteran gut over the timescale of passage of food through the gut, with 50% of trypsin and 40% of chymotrypsin activity remaining after 6h at pH 12, 37 degrees C. Soybean Kunitz trypsin inhibitor (SKTI) ingestion by the larvae had a growth-inhibitory effect, and induced inhibitor-insensitive trypsin-like activity. Qualitative and quantitative changes in proteinase activity bands after gel electrophoresis of gut extracts were evident in SKTI-fed larvae when compared with controls, with increases in levels of most bands, appearance of new bands, and a decrease in the major proteinase band present in extracts from control insects.     

Development and evaluation of an enzyme-linked immunosorbent assay to detect Pieris rapae remains in guts of arthropod predators

An enzyme-linked immunosorbent assay (ELISA) was developed to detect remains of Pieris rapae L. (Lepidoptera: Pieridae) immature stages in the guts of field collected arthropod predators. The assay can be used to help ascertain the relative importance of arthropod predator species in suppressing P. rapae in cabbage, Brassica oleracea var. capitata L. The ELISA is sensitive to all immature stages of P. rapae, although first and fifth instars can be detected more readily than eggs or pupae and third instars showed intermediate detectability. Assays on whole body homogenates of predators readily detected predation on P. rapae first instars by all seven of the predator species tested, although response generally declined with increasing predator size. Together the results show that the P. rapae ELISA possesses a sufficiently high level of sensitivity and specificity to be a useful tool in helping to elucidate the roles of arthropod predator species in reducing populations of P. rapae in cabbage.     

Host plant and the predator Podisus nigrispinus: when the defense compounds of the plant affect the third trophic level

Many herbivorous insects can overcome chemical plant defenses, using the plant's defensive products for their own good, as a defense against predators. Eucalyptus spp. (Myrtaceae), recently introduced in Brazil, are rich in secondary compounds; however, there are reports that these plants have been suffering from population outbreaks of defoliating Lepidoptera in Brazil. The predator Podisus nigrispinus (Dallas) (Hemiptera: Pentatomidae) has been used against herbivorous insects in eucalyptus plantations, but little is known about its establishment in the field. This study aims to investigate whether the effectiveness of this predator may be affected indirectly by compounds of eucalyptus plants, when compared to guava, Psidium guajava L., a Brazilian native species of Myrtaceae. Thus, we evaluated the performance of P. nigrispinus on larvae of Thyrinteina arnobia (Stoll) (Lepidoptera: Geometridae) reared on eucalyptus (exotic species) or guava plants (native species). Podisus nigrispinus performance (reproduction and survival) was better on larvae fed on guava than on larvae fed on eucalyptus. It is possible that the negative effect on the predator's development occurred because of the plants’ secondary compounds appropriated by caterpillars, due to the short coevolutionary history between eucalyptus and the predator. The data suggest that the chemical compounds that could help the plant's defenses against herbivores may also affect their natural enemies, especially when the interaction between plant and natural enemy involves an exotic plant recently introduced into the insect's habitat.