Effects of snowdrop lectin (GNA) delivered via artificial diet and transgenic plants on the development of tomato moth (Lacanobia oleracea) larvae in laboratory and glasshouse trials

The effects of snowdrop lectin (Galanthus nivalis agglutinin, GNA) on Lacanobia oleracea larval growth, development, consumption, and survival, were examined by 3 distinct bioassay methods. Larvae were reared on artificial diet containing GNA at 2% (w/w) dietary protein; on excised leaves of transgenic potato expressing GNA at approx. 0.07% of total soluble proteins; and on transgenic potato plants expressing GNA at approx. 0.6% of total soluble proteins in glasshouse trials. Significant effects on larval growth were observed with all three treatments. At 21days after hatch mean larval biomass was reduced by 32 and 23%, in the artificial diet and excised leaf bioassays respectively. In glasshouse trials a 48% reduction in insect biomass per plant was observed after 35days. The artificial diet and excised leaf assays also showed that GNA significantly slowed larval development as assessed by instar duration. GNA caused a 59% overall reduction in mean daily consumption in the artificial diet assay, and a significant reduction in leaf damage in glasshouse trials. However, prolonged compensatory feeding by larvae in the excised leaf assay resulted in their consuming 15% more total leaf material than the control group. Adaptation to low levels of GNA, in terms of biomass recovery and compensatory feeding, was observed within one larval generation in the detached leaf assay. No significant effects of GNA on larval survival were observed in the artificial diet and detached leaf bioassays, whereas survival was decreased by approx. 40% in the glasshouse bioassay. The assays show that the insecticidal effects of GNA can be observed both in vitro when fed in artificial diet and in planta, and can be demonstrated in the glasshouse as well as under growth cabinet conditions.     

The ectoparasitic wasp Eulophus pennicornis (Hymenoptera: Eulophidae) uses instar-specific endocrine disruption strategies to suppress the development of its host Lacanobia oleracea (Lepidoptera: Noctuidae)

To successfully complete its development, the gregarious ectoparasitoid Eulophus pennicornis must inhibit the moult of its host, Lacanobia oleracea. In the present study, we examined the possibility that moult- and metamorphosis-associated endocrine events may be disrupted in caterpillars parasitized as newly moulted last (sixth) instars. Juvenile hormone (JH) titres on days 2 and 5 of the final stadium were significantly higher (> 100 fold) in parasitized than in non-parasitized hosts, in which JH was essentially absent. Elevated JH levels were associated with reduced haemolymph JH esterase (JHE) activity (down by 99.8%) and enhanced in vitro JH biosynthesis by the corpora allata (CA) (up to 4.5 fold). Wasp adults and/or larvae, in which we measured high levels of JH III (up to 2.7 ng/g), but little or no JH I or JH II, were not seen as likely sources of JH in parasitized hosts, in which we found mostly JH I and JH II. In addition, removal of parasitoid eggs or larvae after oviposition did not prevent the rise in JH titres seen in parasitoid-laden hosts, suggesting that wasp venom may be responsible for the observed hormonal dysfunction. Host haemolymph 20-hydroxyecdysone (20-E) levels were largely unaffected by parasitism during the final stadium although they were observed to increase earlier and decrease more rapidly in parasitized insects. We compare these results with those reported earlier for L. oleracea larvae parasitized by E. pennicornis as penultimate (fifth) instars, which display significantly depressed 20-E titres relative to control larvae. We conclude that E. pennicornis employs host endocrine-disruption strategies that differ according to whether the host is parasitized as a penultimate or final-stadium larva.     

Transgenic GNA Expressing Potato Plants Augment the Beneficial Biocontrol of Lacanobia Oleracea (Lepidoptera: Noctuidae) by the parasitoid Eulophus Pennicornis (Hymenoptera; Eulophidae)

The effect of expressing the gene encoding snowdrop lectin (Galanthus nivalis agglutinin, GNA) in transgenic potato plants, on parasitism of the phytophagous insect pest Lacanobia oleracea by the gregarious ectoparasitoid Eulophus pennicornis, was investigated in glasshouse trials. Expression of GNA (approx. 1.0% total soluble protein) by transgenic plants significantly reduced the level of pest damage, thus confirming previous studies. Furthermore, the presence of the parasitoid significantly reduced the levels of damage incurred either by the transgenic or control plants when compared to those plants grown in the absence of the parasitoid. For the GNA expressing plants the presence of the parasitoid resulted in further reductions (ca. 21%) in the level of damage caused by the pest species. The ability of the wasp to parasitise and subsequently develop on the pest larvae was not altered by the presence of GNA in the diet of the host. E. pennicornis progeny that developed on L. oleracea reared on GNA expressing plants showed no significant alteration in fecundity when compared with wasps that had developed on hosts fed on control potato plants, although mean size and longevity of female parasitoids was significantly reduced. The number of F ₂ progeny produced by parasitoids derived from hosts fed on GNA-expressing plants was not significantly different to those produced by parasitoids from hosts fed control plants. Results from the present study demonstrate that the use of transgenic plants expressing insecticidal proteins can be compatible with the deployment of beneficial insects and that the two factors may interact in a positive manner.     

The snowdrop lectin Galanthus nivalis agglutinin (GNA) and a fusion protein ButaIT/GNA have a differential affect on a pest noctuid Lacanobia oleracea and the ectoparasitoid Eulophus pennicornis

Fusion proteins have considerable potential as novel insect control agents because they enable the oral delivery of insecticidal peptides to the haemolymph of pests. Transport is achieved via fusion of the toxin to a carrier protein Galanthus nivalis agglutinin (GNA) that, after ingestion, binds to and crosses the insect gut epithelia. A fusion protein comprising a toxin from the South Indian red scorpion (Mesobuthus tamulus) that is fused to a GNA polypeptide (ButaIT/GNA) has a detrimental effect on the development of tomato moth Lacanobia oleracea (L.) (Lepidoptera: Noctuidae) larvae. The present study examines the effects of ButaIT/GNA and GNA, delivered orally or by injection, on the development of L. oleracea larvae, and the subsequent effects on the gregarious ectoparasitoid Eulophus pennicornis (Nees) (Hymenoptera: Eulophidae) developing on ButaIT/GNA‐ and GNA‐treated hosts. The fusion protein, but not GNA, reduces the growth of fifth stadium L. oleracea larvae. The development of E. pennicornis is not affected by the presence of ButaIT/GNA in hosts that ingest the protein, although it is affected when hosts are injected with the protein. This difference is considered to be a result of higher levels of fusion protein being present when the fusion protein is injected. Intact ButaIT/GNA is detected by immunoassay in the haemolymph of L. oleracea larvae after ingestion of the fusion protein. More unexpectedly, negative effects are observed for the growth of E. pennicornis larvae developing on hosts that have either ingested, or been injected with GNA.     

The biology of Meteorus gyrator (Hymenoptera: Braconidae), a solitary endoparasitoid of the tomato moth, Lacanobia oleracea (Lepidoptera: Noctuidae)

There is a need to identify potential biological control agents for use against lepidopterous pests in greenhouses. The solitary endoparasitoid Meteorus gyrator (Thunberg) attacks a range of macrolepidopterous larvae, including those of some important horticultural pest species. Laboratory trials designed to investigate the biology of M. gyrator on larvae of the tomato moth, Lacanobia oleracea Linnaeus, reveal that this parasitoid is capable of parasitizing all larval stages of its host, third instars being parasitized most frequently. Each female parasitoid lives for up to 40 days (at 25 degrees C), ovipositing into an average of 78 hosts. Preadult development is rapid ( approximately 2 weeks), and the sex ratio of offspring is 1:1. Parasitism by M. gyrator suppresses the growth of both early and late host instars, and there is a concomitant reduction in the amount of food consumed (overall feeding reduction over a 12 day period is 68%). Our results indicate that inoculative releases of M. gyrator could provide effective biological control of L. oleracea and other noctuid pests of greenhouses.     

Effects of Galanthus nivalis agglutinin (GNA) expressed in tomato leaves on larvae of the tomato moth Lacanobia oleracea (Lepidoptera: Noctuidae) and the effect of GNA on the development of the endoparasitoid Meteorus gyrator (Hymenoptera: Braconidae)

The effect of ingestion of transgenic tomato leaves expressing the plant lectin Galanthus nivalis agglutinin (GNA) on development of larvae of Lacanobia oleracea (Linnaeus) was studied under laboratory conditions. When L. oleracea larvae were fed on tomato line 14.1H, expressing approximately 2.0% GNA, significant increases in the mean larval weight and in the amount of food consumed were found. This resulted in an overall reduction in the mean development time to the pupal stage of approximately 7 days. A significant increase in the percentage survival to the adult moth was also recorded when newly hatched larvae were reared on transgenic tomato leaves (72%) compared to larvae reared on untransformed leaves (40%). The effects of ingestion of GNA by L. oleracea larvae, via artificial diet or the leaves of transgenic tomato or potato plants, on the subsequent development of its solitary endoparasitoid Meteorus gyrator (Thunberg) was also studied. No significant effects on the life cycle parameters of M. gyrator developing in L. oleracea fed on GNA-containing diets were observed. Experiments with transgenic potato plants indicated that the stadium of the host larvae at parasitism had a greater influence on M. gyrator development than the presence of GNA. Potential GNA-binding glycoproteins were detected in the gut and body tissues of larval M. gyrator. Despite detection in host tissues, GNA could not be detected in adult M. gyrator and therefore it is likely that at the time of pupation M. gyrator are able to void the GNA in the meconial pellet.     

Longevity and fecundity of Eulophus pennicornis, an ectoparasitoid of the tomato moth Lacanobia oleracea, is affected by nutritional state and diet quality

• 1 Adult female Eulophus pennicornis require a source of nutrition, provided by sources such as pollen, nectar and honeydew or by host feeding, to promote longevity and facilitate egg production. There is potential for parasitoids to be exposed directly to contaminants, including gene products in transgenic crops, through feeding on plant materials, honeydew or hosts. Among such potential contaminants are lectins such as Galanthus nivalis agglutinin (GNA) and concanavalin agglutinin (Con A).
• 2 The effect of direct exposure to honey diets containing GNA and Con A on the longevity and fecundity of E. pennicornis was examined. These lectins have been expressed in a number of plant species for the control for various insect pests. Both GNA and Con A significantly reduced longevity and fecundity at the highest concentration used (0.5% w/v). The effect on fecundity was shown to be related to a reduction in longevity.
• 3 Examination of the gustatory response of adult female E. pennicornis to honey diet containing 1% w/v GNA or Con A revealed no significant differences in consumption rate on first exposure. A significant reduction in the time spent feeding on diet containing 1% Con A was found on the second exposure to the diet. This could have been the result of either a conditioned aversion response or the intoxication of the insect. The effect of Con A on longevity and fecundity could have been, in part, a result of reduced food intake.
• 4 Studies on nutrition and egg resorption demonstrated that the availability of honey solution prolongs the longevity of E. pennicornis and the lack of a source of nutrition promotes oosorption.
• 5 A greater understanding of feeding behaviour and ovigeny is required to understand fully the potential ecological consequence of transgenic crops on parasitoid species through routes of direct exposure to transgene products.

     

Parasitization of Lacanobia oleracea (Lepidoptera: Noctuidae) by the ectoparasitc wasp,Eulophus pennicornis. Effects of parasitization,venom and starvation on host haemocytes

In contrast to the situation with endoparasitic wasps, little is known about the effects of ectoparasitoids and their secretions on the haemocytes of their insect hosts. To address this deficit, a study has been made of the ectoparasitic wasp, Eulophus pennicornis, and it's host, the tomato moth, Lacanobia oleracea. Using light microscopy, it was determined that L. oleracea has five main haemocyte types, namely, plasmatocytes, granular cells, spherule cells, oenocytoids and pro-haemocytes, representing 56%, 30%, 10%, 2% and 2% of the population, respectively. Parasitization by E. pennicornis, resulted in an increase in the number of circulating haemocytes up to day three, followed by a decrease towards day eight; the latter being associated with changes to the morphology and viability of the cells. For example, on day five after parasitization, plasmatocytes and granular cells had become more rounded and put out pseudopods less readily compared with those from non-parasitized controls, whilst from day seven onwards there was a significant decrease in haemocyte viability and by day nine, extensive haemocyte damage and disintegration was evident. These changes were not observed when larvae were injected with E. pennicornis venom, or when haemocytes were exposed directly to venom in vitro, neither did they occur in starved larvae. Thus, although the observed effects on L. oleracea haemocytes are definitely associated with parasitization they are not due to wasp venom components, nor are they a non-specific effect resulting from nutritional deprivation. The possibility that the feeding wasp larvae produce factors which perturb host haemocytes in order to help condition the host to ensure that successful parasitization occurs, is discussed.     

The effects of Phaseolus vulgaris erythro- and leucoagglutinating isolectins (PHA-E and PHA-L) delivered via artificial diet and transgenic plants on the growth and development of tomato moth (Lacanobia oleracea) larvae; lectin binding to gut glycoproteins in vitro and in vivo

Red kidney bean, Phaseolus vulgaris, contains a lectin phytohemagglutinin (PHA) with toxicity towards higher animals. PHA exists in the isoforms PHA-E and PHA-L, which agglutinate erythrocytes and lymphocytes, respectively. Lacanobia oleracea larvae were reared from hatch on artificial diets containing PHA-E or PHA-L at 2% (w/w) dietary protein, and on transgenic Arabidopsis plants expressing either lectin at 0.4-0.6% of total soluble proteins. In artificial diet bioassays neither lectin affected larval survival, development, growth nor consumption. In transgenic plant bioassays both PHA-E and PHA-L promoted larval growth and development. This effect was greatest for PHA-E. Mean larval biomass of insects fed on plants expressing PHA-E was significantly greater (up to two-fold) than controls during the final two instars and the insects developed at a significantly greater rate so that after 26 days 83% of PHA-E exposed insects were in the final instar compared to 44% for control insects. PHA-E and PHA-L were detected by Western blotting in haemolymph, sampled from insects fed diets or plant material containing the lectins. However, despite the demonstrated potential for both isolectins to bind to gut glycopolypeptides in vitro neither was found to accumulate in vivo in the guts of exposed insects. Since lectin binding to gut polypeptides is thought to be necessary for insecticidal activity the failure of PHA-E and PHA-L to bind in vivo may account for their lack of toxicity to L. oleracea.     

Neochrysocharis formosa (Westwood) (Hymenoptera: Eulophidae), a newly recorded parasitoid of the tomato moth, Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae), in Argentina

We report the first record of Neochrysocharis formosa (Westwood) parasitizing larvae of the tomato moth, Tuta absoluta (Meyrick), in tomato crops in Northern Buenos Aires Province, Argentina. Tomato moth larvae were sampled during four consecutive growing cycles, between 2003 and 2005, in 10 sites. Neochrysocharis formosa was present only in organic outdoor and protected crops, and predominantly during the late season. Parasitism rates varied from 1.5% to 5%. The finding of this species is a new record for Argentina and South America, and T. absoluta is a new host record.     

Parasitization of Lacanobia oleracea (Lepidoptera) by the ectoparasitic wasp,Eulophus pennicornis,suppresses haemocyte-mediated recognition of non-self and phagocytosis

Although many endoparasitic wasps suppress the haemocyte-mediated immune defences of their insect hosts, the effects of ectoparasitoids are virtually unknown. In view of this, a study has been made of the ectoparasitic wasp, Eulophus pennicornis, and its host, the tomato moth, Lacanobia oleracea. For unparasitized insects, in vitro assays indicated that less than 3.0% of L. oleracea haemocytes on a monolayer formed rosettes with yeast cells or fresh rabbit erythrocytes (rbc), and virtually no phagocytosis of these particles occurred. In addition, although fixed rbc formed rosettes with 51.21% of haemocytes, only about 3.0% of the haemocytes ingested one or more of these particles. In contrast to this, B. cereus and E. coli were readily phagocytosed by 14.75% and 53.70% of haemocytes, respectively. These results indicate that L. oleracea haemocytes can recognise different types of non-self particles and demonstrate that ingestion does not necessarily follow attachment. When rosetting and phagocytosis assays were performed with fixed rbc and FITC-labelled E. coli, and haemocytes from starved L. oleracea, PBS injected L. oleracea, and experimentally envenomated insects on day five of treatment, there was no significant difference in the percentage of rosetting or phagocytosis occurring. When haemocytes from parasitized insects on day five of treatment were utilised, however, rosetting and phagocytosis were reduced by 31.41% and 34.94%, respectively. Thus, the effects of parasitization and experimental envenomation are not the same. In addition, suppression of host haemocyte-mediated recognition and phagocytosis was not a secondary effect of nutritional deprivation and was not due to ectoparasitoid venom components, rather it was a direct result of parasitization of L. oleracea by E. pennicornis. The putative nature and source of the immunosuppressive factor(s) involved is discussed with reference to those produced by endoparasitic wasps.     

Parasitism of Lacanobia oleracea (Lepidoptera) by the ectoparasitoid, Eulophus pennicornis, is associated with a reduction in host haemolymph phenoloxidase activity

When haemolymph from fifth instar Lacanobia oleracea was incubated in vitro, rapid melanization occurred. Similar levels of melanization occurred in haemolymph from larvae that had been experimentally injected with venom from the ectoparasitic wasp, Eulophus pennicornis. In contrast, haemolymph from larvae parasitized by this wasp melanized more slowly and less extensively. Phenoloxidase assays indicated that enzyme activity was present in haemocyte lysate supernatants, serum and plasma from L. oleracea and that on day 5 post-parasitization, fractions prepared from parasitized larvae had significantly less phenoloxidase activity than similar fractions from untreated or experimentally envenomated larvae. In addition, no PO activity was detectable in wasp venom, and the venom had no effect on L. oleracea plasma phenoloxidase activity in vitro. These results indicate that parasitism of L. oleracea by E. pennicornis suppresses host haemolymph phenoloxidase activity and that this suppression is not induced by adult wasp venom. The results are discussed with reference to the survival advantages of suppressing the activity of this host enzyme, and to the possible source(s) of putative suppressive factors.     

Expression of snowdrop lectin (GNA) in transgenic rice plants confers resistance to rice brown planthopper

Snowdrop lectin ( Galanthus nivalis agglutinin; GNA) has been shown previously to be toxic towards rice brown planthopper ( Nilaparvata lugens ; BPH) when administered in artificial diet. BPH feeds by phloem abstraction, and causes ‘hopper burn’, as well as being an important virus vector. To evaluate the potential of the gna gene to confer resistance towards BPH, transgenic rice ( Oryza sativa L.) plants were produced, containing the gna gene in constructs where its expression was driven by a phloem-specific promoter (from the rice sucrose synthase RSs1 gene) and by a constitutive promoter (from the maize ubiquitin ubi1 gene). PCR and Southern analyses on DNA from these plants confirmed their transgenic status, and that the transgenes were transmitted to progeny after self-fertilization. Western blot analyses revealed expression of GNA at levels of up to 2.0% of total protein in some of the transgenic plants. GNA expression driven by the RSs1 promoter was tissue-specific, as shown by immunohistochemical localization of the protein in the non-lignified vascular tissue of transgenic plants. Insect bioassays and feeding studies showed that GNA expressed in the transgenic rice plants decreased survival and overall fecundity (production of offspring) of the insects, retarded insect development, and had a deterrent effect on BPH feeding. gna is the first transgene to exhibit insecticidal activity towards sap-sucking insects in an important cereal crop plant.     

Oral toxicity and impact on fecundity of three insecticidal proteins on the gregarious ectoparasitoid Eulophus pennicornis (Hymenoptera: Eulophidae)

Abstract 1 The effects of three insecticidal transgene proteins on selected life parameters of the ectoparasitoid Eulophus pennicornis were investigated. 2 When incorporated into the diet of the adult wasp, the lectins GNA (snowdrop lectin) and Con A (jackbean lectin) significantly reduced longevity at doses of 0.1% w/v and above. At a dose rate of 0.1% w/v, GNA and Con A reduced mean longevity to approximately 13 and 10 days, respectively, compared with average control lifespans of approximately 20 days. The trypsin inhibitor from cowpeas, CpTI, had no marked effect on longevity. 3 Both lectins reduced reproductive fitness of parasitoids when dosed before exposure to hosts. The 1.0% dose reduced fecundity by over 35% for GNA and 70% for Con A. Although reduced fecundity may have been a function of shorter lifespans, smaller egg loads in female wasps provided evidence to suggest that higher lectin doses may have interfered with egg maturation processes. 4 Both lectins were readily detected in whole body extracts of the parasitoid and were seen to persist within their bodies for at least 24 h after cessation of feeding on lectin‐containing diets. 5 The results would indicate that the ingestion of the lectins, either through host feeding or through the consumption of nectar, may pose a hazard to parasitic wasps if present at sufficiently high concentrations.     

Infection by the microsporidium Vairimorpha necatrix (Microspora: Microsporidia) elevates juvenile hormone titres in larvae of the tomato moth, Lacanobia oleracea (Lepidoptera: Noctuidae)

The effects of infection by a microsporidium, Vairimorpha necatrix (Kramer), on the endogenous levels of juvenile hormones in tomato moth (Lacanobia oleracea L.) larvae were investigated. Levels of juvenile hormone II (JH II) were 10-fold greater in the infected larvae on day two of the sixth stadium but no significant difference was observed on day seven. Juvenile hormone I (JH I) was also detected in day two and day seven sixth instar infected larvae but was not detected in non-infected larvae. The duration of the fifth and sixth stadia was significantly longer for infected larvae when compared with non-infected larvae. No evidence was found to suggest that supernumerary moults are a feature of infection by V. necatrix in L. oleracea larvae. Experiments were performed to determine whether the elevation in JH levels, which probably prevents pupation, is an adaptive mechanism of the microsporidium for extending the growth phase of the host, thereby allowing increased spore production. A proportion of infected larvae were collected on days 9 and 24 of the sixth stadium and spore extracts prepared from each larva. These days represent the average duration of the sixth stadium required for uninfected larvae to reach pupation, and the average number of days that V. necatrix-infected larvae survive in the sixth stadium before dying from infection. The mean spore yields from infected larvae 24 days into the sixth stadium were significantly higher than the spore yields obtained from day nine sixth instar larvae. The hypothesis that V. necatrix manipulates host endocrinology (i.e. prolong the host larval state to maximise spore yield) is discussed in context with the results obtained.     

Concanavalin A inhibits development of tomato moth (Lacanobia oleracea) and peach-potato aphid (Myzus persicae) when expressed in transgenic potato plants

The effects of concanavalin A (ConA), a glucose/mannose-specific lectin from jackbean (Canavalia ensiformis), on insect crop pests from two different orders, Lepidoptera and Homoptera, were investigated. When fed to larvae of tomato moth (Lacanobia oleracea) at a range of concentrations (0.02–2.0% of total protein) in artificial diet, ConA decreased survival, with up to 90% mortality observed at the highest dose level, and retarded development, but had only a small effect on larval weight. When fed to peach-potato aphids (Myzus persicae) at a range of concentrations (1–9μM) in liquid artificial diet, ConA reduced aphid size by up to 30%, retarded development to maturity, and reduced fecundity (production of offspring) by >35%, but had little effect on survival. With both insects, there was a poor correlation between lectin dose and the quantitative effect. Constitutive expression of ConA in transgenic potatoes driven by the CaMV 35S promoter resulted in the protein accumulating to levels lower than predicted, possibly due to potato not being able to adequately reproduce the post-translational processing of this lectin which occurs in jackbean. However, the expressed lectin was functionally active as a haemagglutinin. Bioassay of L. oleracea larvae on ConA-expressing potato plants showed that the lectin retarded larval development, and decreased larval weights by >45%, but had no significant effect on survival. It also decreased consumption of plant tissue by the larvae. In agreement with the diet bioassay results, ConA-expressing potatoes decreased the fecundity of M. persicae by up to 45%. ConA thus has potential as a protective agent against insect pests in transgenic crops. This revised version was published online in June 2006 with corrections to the Cover Date.     

Development of a gregarious ectoparasitoid, Euplectrus separatae (Hymenoptera; Hymenoptera: Eulophidae), that parasitizes Pseudaletia separata (Lepidoptera: Noctuidae)

Euplectrus separatae is a gregarious ectoparasitoid that parasitizes Pseudaletia separata during its third to sixth (last) instars. The eggs of the parasitoid are fixed on the integument of the host dorsolaterally with a hard substance like a piling driven into the integument by the female wasp at the time of oviposition. The first instar of the parasitoid, which hatches three days after oviposition is nourished by ingesting the hemolymph of the host, and ecdyses to the second stadium six days after oviposition. Many hemocytes and epidermal cells were found assembled under the piling and places where a parasitoid had attached its mouth, suggesting that the host had repaired the integument destroyed by the parasitoid. Botryoidal tissue, which stained well with hematoxylin, began to develop from four days after oviposition and became gradually larger with development. Botryoidal tissue appears to function as a secretory organ for thread and a storage organ for nutrients. Seven days after oviposition, the parasitoid larvae migrate down from the dorsal surface to the ventral side of the host. Just before descending they ecdyse to the third stadium and kill the host during their migration. If all parasitoid larvae were removed artificially from the host before they migrate, the host did not die. However, removing the parasitoids after they had started to migrate did not prevent the death of the host. Transmission electron microscopic (TEM) observation of salivary glands of a parasitoid larva before migrating revealed that the salivary gland was composed of cells that were rich in rough surfaced endoplasmic reticulum (rough-ER) with many ribosomes and cells that were filled with a lot of vacuoles just before their collapse. After moving from the host body, the parasitoid larvae doubled in weight by ingesting the tissue of the host and then spun a cocoon. Almost all host tissues were consumed for growth of the parasitoid, like an idiobiont parasitoid.     

Parasitism of Lacanobia oleracea (Lepidoptera,noctuidae) by the ectoparasitic wasp Eulophus pennicornis,results in the appearance of a 27 kDa parasitism-specific protein in host plasma

Little is known about the effects of ectoparasitoids and their secretions on the plasma protein profiles of their insect hosts. To address this deficit, a study has been made of the interactions between an ectoparasitic wasp, Eulophus pennicornis, and its host, the tomato moth, Lacanobia oleracea. In particular, the quantitative and qualitative effects of parasitism or the experimental injection of wasp venom on host plasma proteins were investigated. Results demonstrated that both treatments caused an initial increase in L. oleracea total plasma protein concentration up to day 5 of treatment, but whereas the protein concentration remained high in the experimentally envenomated group, a decrease towards day 8 occurred in parasitized insects. Parasitism was also associated with the appearance of a protein with an estimated molecular weight of 27 kDa. This protein first appeared on day 3 after parasitization and its levels subsequently increased. The protein was not detected in any of the unparasitized larvae (including all the various control groups) or in experimentally envenomated L. oleracea larvae. In addition, the appearance of this protein was not a non-specific result of nutritional deprivation, nor was it a general injury, stress, or infection induced protein. Its appearance was strictly associated with parasitism of L. oleracea by E. pennicornis and thus, it may be described as a parasitism-specific protein (PSP). The PSP has been partially purified using whole gel elution. Gel filtration and SDS PAGE indicated that it has a native molecular weight of 27 kDa and that it does not appear to aggregate to produce higher molecular weight molecules, nor dissociate into lower molecular weight subunits held together by disulphide or covalent bonds. The precise site of synthesis of the 27 kDa PSP is not yet known but some evidence leads us to speculate that it may be synthesised by the feeding E. pennicornis larvae and introduced into their host. This possibility is discussed in relation to previous work detailing the effects of parasitism on L. oleracea haemocyte morphology, function and viability, and the effects of endoparasitoids on host plasma proteins.     

Fecundity and development of Cirrospilus coachellae (Hymenoptera: Eulophidae), a parasitoid of Marmara gulosa (Lepidoptera: Gracillariidae)

The biology of Cirrospilus coachellae Gates (Hymenoptera: Eulophidae), an ecto-parasitoid of the larvae of Marmara gulosa Guillén & Davis (Lepidoptera: Gracillariidae) was studied in the laboratory. In total, 120.3 +/- 21.2 adults were produced per female at 26 degrees C. Females survived an average of 23.5 +/- 4.4 d and parasitized an average of 49.8 +/- 10.2 hosts over their lifetime. C. coachellae is a facultatively gregarious parasitoid with up to eight parasitoids developing per host with an average of 2.5 +/- 0.1 in the laboratory. A female-biased sex ratio of 0.31 (male/total) was exhibited. Development times decreased with increasing temperatures from 23.7 +/- 0.2 d at 20 degrees C to 9.2 +/- 0.1 at 29 degrees C. Estimated lower and upper development thresholds were 14.1 and 36.7 degrees C, respectively. Maximal development rate occurred at 31.3 degrees C, and 138 degree-days was required to complete the cycle.     

Ability to host regulate determines host choice and reproductive success in the gregarious ectoparasitoid Eulophus pennicornis (Hymenoptera: Eulophidae)

Abstract.  The age of Lacanobia oleracea (L.) in the final (sixth) larval stadium influences host choice and developmental success significantly in the gregarious ectoparasitoid Eulophus pennicornis (Nees). In choice tests, parasitoids with prior oviposition experience parasitize hosts in the second day of the sixth stadium most frequently. Parasitoid brood survival on normally-reared (i.e. fed) hosts declines monotonically with age such that mean progeny survival (egg–adult) is less than 20% for wasps developing on hosts parasitized on day 5 of the sixth stadium, as opposed to almost 50% when developing on those parasitized on day 1. Neck ligation of hosts increases the survival of wasp larvae developing on older hosts (days 4 and 5), whereas starved hosts produce progeny in similar numbers to fed hosts on most days during the final larval stadium. Hosts parasitized early in the stadium (days 1–3), although continuing to grow, do not exhibit the characteristic physical changes that non-parasitized larvae exhibit prior to pupation. However, hosts parasitized on days 4 and 5 form prepupae in appreciable numbers, particularly on day 5 where, regardless of treatment, over 80% of hosts attain this stage. Envenomated hosts behave similarly, an observation that suggests that it is the wasp's inability to arrest completely development in older hosts that is the significant factor in reducing the developmental success of the wasp. The findings are discussed in the light of the known endocrinological events in the host, and in relation to previously reported host manipulations induced by this wasp.