Tolerance to Bacillus thuringiensis endotoxin in immune-suppressed larvae of the flour moth Ephestia kuehniella.

Tolerance to Bacillus thuringiensis crystal endotoxins (Bt-toxins) is correlated with an elevated immune status in larvae of the flour moth Ephestia kuehniella. To gain more specific information about the effector pathways involved in the protection against the toxin, we studied the effects of Bt-toxin formulations in susceptible (non-induced) and tolerant (immune-induced) larvae after natural (parasitism-mediated) and chemical (tropolone-mediated) suppression of defence reactions. Although melanization in hemolymph was significantly reduced, there was no significant effect on susceptibility to the toxin in parasitised or tropolone-treated larvae. This suggests that melanization of hemolymph is correlated with an elevated immune status but not responsible for the observed tolerance to Bt-toxin. To examine whether hemolymph proteins exist in the gut lumen and function as pro-coagulants, we compared gut and plasma proteins of immune-induced with those of non-induced larvae. Here we show that the lipid carrier lipophorin represents a major component in the gut lumen and interacts with mature Bt-toxin to form a complex.

Factors affecting growth in the koinobiont endoparasitoid Venturia canescens in the flour moth Ephestia kuehniella.

With resistance of insect pests to synthetic pesticides on the increase, the role of parasitoid wasps as biological control agents is expanding in pest and resistance management strategies. One of the predictors of reproductive success of endoparasitoids is the relative size of the wasp at host emergence. While in idiobiont parasitoids, where the host stops feeding after parasitism, the wasp size is determined by the host size at the time of parasitism; the size of koinobiont wasps, where the host continues to feed after parasitism, is dependent on additional factors. Here we show that the host mass and temperature are important factors that determine survival and development of the koinobiont endoparasitoid Venturia canescens in late instar larvae of the flour moth Ephestia kuehniella.

Cell-free immune reactions in insects.

Insects, like many other multicellular organisms, are able to recognise and inactivate potential pathogens and toxins in the absence of cells. Here we show that the recognition and inactivation of lipopolysaccharides (LPS) and bacteria is mediated by lipophorin particles, which are the lipid carrier in insects. In immune-induced insects sub-populations of lipophorin particles are associated with pattern recognition proteins and regulatory proteins that activate prophenoloxidase. Moreover, interactions with lectins result in the assembly of lipophorin particles into cage-like coagulation products, effectively protecting the surrounding tissues and cells from the potentially damaging effects of pathogens and phenoloxidase products. The existence of cell-free defence reactions implies that immune signals exist upstream of cell-bound receptors.

An empirical model of the sympatric coexistence of two strains of the endoparasitoid wasp Venturia canescens.

Recent research has demonstrated that a laboratory culture of the asexual solitary endoparasitoid wasp Venturia canescens Grav. (Hymenoptera: Ichneumonidae) contains two genetically and phenotypically distinct lines, coexisting on their host the flour moth Ephestia kuehniella Zeller (Lepidoptera: Pyralidae). The basis to the coexistence of the two lines appears to be differences in their reproductive success under single parasitism and superparasitism. Furthermore, examination of field-derived wasps from several locations has shown that the phenotypes displayed by the laboratory colonies also co-occur in field populations. Historically, the impossibility of showing that two species do not occupy separate niches has precluded any demonstration of sympatric coexistence in the field. Here we present the results of an iterative model that uses a range of experimental life history data to predict the stable composition of a mixed population of two lines displaying the laboratory phenotypes under different rates of superparasitism. The model predicts that sympatric coexistence of the two lines is possible when the overall rate of superparasitism is between 4 and 12% or greater. These values are within the rates reported for other solitary endoparasitoid wasp species in the field, and so demonstrate that the sympatric coexistence under natural conditions of two species that display the phenotypes observed in the laboratory lines is, in principle, possible.

Are insect immune suppressors driving cellular uptake reactions?

Many insect parasitoids that deposit their eggs inside immature stages of other insect species inactivate the cellular host defence to protect the growing embryo from encapsulation. Suppression of encapsulation by polydnavirus-encoded immune-suppressors correlates with specific alterations in hemocytes, mainly cytoskeletal rearrangements and actin-cytoskeleton breakdown. We have previously shown that the Cotesia rubecula polydnavirus gene product CrV1 causes immune suppression when injected into the host hemocoel. CrV1 is taken up by hemocytes although no receptors have been found to bind the protein. Instead CrV1 uptake depends on dimer formation, which is required for interacting with lipophorin, suggesting a CrV1-lipophorin complex internalisation by hemocytes. Since treatment of hemocytes with oligomeric lectins and cytochalasin D can mimic the effects of CrV1, we propose that some dimeric and oligomeric adhesion molecules are able to cross-link receptors on the cell surface and depolymerise actin by leverage-mediated clearance reactions in the hemolymph.

Phenoloxidase-like activities and the function of virus-like particles in ovaries of the parthenogenetic parasitoid Venturia canescens.

The ichneumonid endoparasitoid Venturia canescens successfully develops inside the hemocoel of another insect by using maternal protein secretions, including nucleic acid-free virus-like particles (VLPs), to manipulate host physiology. These VLPs consist of four major proteins, which are produced mainly in the calyx tissue and transferred into the host insect together with the egg. One of the protein-coding genes (vlp1), with similarities to phospholipid-hydroperoxide glutathione peroxidases (PHGPx), exists in allelic forms producing two protein variants with different protein properties. Here, we summarise observations indicating that oocytes and eggs are the source of reactive electrons, which potentially damage the lining and membranes of calyx tissues. We discuss the possible role of VLP1 in counteracting the damaging effects of oxidised phospholipids on membranes surrounding VLPs in the calyx lumen.

The development of the endoparasitoid Venturia canescens in Bt-tolerant, immune induced larvae of the flour moth Ephestia kuehniella.

We examined the survival and development of the endoparasitoid Venturia canescens in a Bt-tolerant laboratory strain of the flour moth Ephestia kuehniella, in which Bt-tolerance has been shown to be associated with an inducible elevation of the insect's immune response. The results indicate the elevated immune status associated with Bt-tolerance does not confer cross-protection against parasitism by V. canescens. No significant difference was observed in the rate of emergent wasps from parasitised Bt-tolerant and Bt-susceptible hosts. In addition, wasps from Bt-tolerant hosts had longer development times and were larger than wasps from Bt-susceptible hosts.

Lectin-induced haemocyte inactivation in insects.

Most multimeric lectins are adhesion molecules, promoting attachment and spreading on surface glycodeterminants. In addition, some lectins have counter-adhesion properties, detaching already spread cells which then acquire round or spindle-formed cell shapes. Since lectin-mediated adhesion and detachment is observed in haemocyte-like Drosophila cells, which have haemomucin as the major lectin-binding glycoprotein, the two opposite cell behaviours may be the result of lectin-mediated receptor rearrangements on the cell surface. To investigate oligomeric lectins as a possible extracellular driving force affecting cell shape changes, we examined lectin-mediated reactions in lepidopteran haemocytes after cytochalasin D-treatment and observed that while cell-spreading was dependent on F-actin, lectin-uptake was less dependent on F-actin. We propose a model of cell shape changes involving a dynamic balance between adhesion and uptake reactions.

The development of the endoparasitoid wasp Venturia canescens in superparasitised Ephestia kuehniella.

Using a molecular marker that allows the differentiation of two strains of the solitary endoparasitoid wasp Venturia canescens, the study investigated the influence of host mass and the time interval between ovipositions on the survival and development of larvae from both the first and second laid eggs in superparasitised Ephestia kuehniella. As the time interval between ovipositions increased both overall and superparasitism success decreased, however, time between, and order of, ovipositions had little effect on other developmental parameters. Adult size increased with host mass under both parasitism and superparasitism, while host mortality decreased with host mass under superparasitism. In addition, wasps emerging from superparasitised hosts were larger than wasps from parasitised hosts. The results confirm that for V. canescens on the host E. kuehniella both self- and conspecific-superparasitism will be an adaptive strategy when hosts are the limiting factor.

Two coexisting lines of the endoparasitoid Venturia canescens show differences in reproductive success under conspecific superparasitism.

In a laboratory colony of the endoparasitic wasp Venturia canescens Grav. (Hymenoptera: Ichneumonidae), two genetically distinct lines (RP, RM) appear to coexist sympatrically. The two lines display pronounced differences in ovarian morphology, parasitism behaviour and number of offspring produced under competing superparasitism. Since V. canescens is a solitary endoparasitoid, larvae inside superparasitised hosts must compete for host possession. We examined the outcome of conspecific superparasitism between the wasp lines with different time intervals between ovipositions. The results showed that the competitive abilities of the two lines were not symmetrical. Further, the RM-line won a significantly higher fraction (around 60%) of the overall contests. Dissection of parasitoid larvae from their hosts indicated that most contests between competing larvae had occurred within the first 24 h of the eggs hatching, suggesting the advantage of the RM-line relates to physical combat. It was previously thought that the coexistence of the two lines was exclusively due to maternal effects. The results of this study indicate for the first time that these differences are based on phenotypic variations in both the larval offspring and the mother.

Lifetime egg maturation by host-deprived Venturia canescens.

The study investigated egg maturation and deposition by the endoparasitoid wasp Venturia canescens under conditions of host deprivation. Female V. canescens maintained without hosts began to deposit eggs onto the sides of the culture vessel on the day of eclosion. The maturation of additional eggs was not inhibited once the maximum oviduct egg load was reached but rather continued for the duration of the experiment (up to 39 days), at a rate of around 5.8% of the remaining unmatured eggs per day. Following host access, wasps matured additional eggs at an increased rate. Artificial damage to the ovipositor resulted in a reduced rate of egg maturation even though the oviducts were partly egg depleted, while damage to the auxiliary valvulae had no effect. These results suggest two conclusions. Under conditions of host deprivation, the rate at which eggs are matured is determined by the rate of synthesis of precursors by the fat body that in turn is modified by feedback from the ovipositor, induced by physical stimulation. Further, the discarding of eggs is due to the involuntary unidirectional movement of eggs down the oviduct, facilitated by the ongoing maturation of additional eggs.

Induction and transmission of Bacillus thuringiensis tolerance in the flour moth Ephestia kuehniella.

The use of Bacillus thuringiensis (Bt) endotoxins to control insect vectors of human diseases and agricultural pests is threatened by the possible evolution of resistance in major pest species. In addition to high levels of resistance produced by receptor insensitivity (5, 16, 17), several cases of tolerance to low to medium levels of toxin have been reported in laboratory colonies of lepidopteran species (3, 18). Because the molecular basis of some of these cases of tolerance to the toxin are not known, we explored alternative mechanisms. Here, we present evidence that tolerance to a Bt formulation in a laboratory colony of the flour moth Ephestia kuehniella can be induced by preexposure to a low concentration of the Bt formulation and that the tolerance correlates with an elevated immune response. The data also indicate that both immune induction and Bt tolerance can be transmitted to offspring by a maternal effect and that their magnitudes are determined by more than one gene.