Nonparasitized Larvae Research Articles

Inhibition of testicular growth and development in Manduca sexta larvae parasitized by the braconid wasp Cotesia congregata

Tobacco hornworm larvae parasitized by the gregarious larval endoparasitoid Cotesia congregata exhibited an inhibition in testicular growth and development, the extent of which was determined by the age and developmental stage of the host at the time of parasitization. The degree of parasitic castration, as assessed by measurements of testicular volume, was correlated with the stadium in which parasitization occurred. A mathematical formula requiring the measurement of testicular length, width and depth was used to calculate testicular volume. The use of the depth parameter revealed a negative correlation between host weight and testicular volume in parasitized larvae. Testicular volumes of fifth instar hosts, which had been parasitized in the first stadium, were significantly smaller than those originally parasitized as fourth or fifth instar larvae and were not correlated with parasitoid load. Effects of natural parasitism were not duplicated by injections of C. congregata polydnavirus and venom, topical treatment with the juvenile hormone analog methoprene, or starvation of nonparasitized larvae. Larvae receiving virus plus venom or methoprene grew larger due to delayed wandering and had larger testes than controls. Deleterious effects on host testes may be due to the effects of nutrient competition between the developing parasitoid progeny and the gonads, combined with the juvenilizing effects believed to be caused by the polydnavirus.

Inhibitory effects of parasitism by the gregarious endoparasitoidCotesia congregata on host testicular development

Cotesia congregata is a gregarious larval endoparasitoid of the tobacco hornworm, Manduca sexta. Parasitized larvae exhibit a variety of physiological and developmental aberrations, the most obvious of which is the induction of developmental arrest characterized by the absence of wandering behavior and suppression of pupation. This arrest appears attributable to continued maintenance of an elevated titer of juvenile hormone and reduced levels of hemolymph juvenile hormone esterase activity. Injection of the wasp's polydnavirus into nonparasitized larvae also causes arrest and the larvae eventually form larval-pupal intermediates instead of normal pupae, indicating the virus may be partially responsible. Aside from causing arrested host development, parasitism also inhibits the normal development and differentiation of testes in male host larvae, so that the testes atrophy instead of growing synchronously with other larval tissues. Here we report that parasitism has pronounced disruptive cytological effects on the developing reproductive organs of male hosts, in addition to causing them to atrophy. Parasitism results in a reduction in testicular volume attributable to a reduction in the number of developing germ cells. Microscopy revealed that the structural integrity of the sheaths surrounding the testicular follicles also is disrupted, so that the tissues appear grossly abnormal compared to those of nonparasitized larvae. Intrahemocoelic injection of purified C. congregata polydnavirus in combination with venom into nonparasitized fourth instar larvae, or topical application of 100 μg of methoprene to fourth instar larvae, also alters sheath integrity and reduces the numbers of developing germ cells, but not to the same degree as the pattern observed in truly parasitized hosts. The occurrence of cell death in the male gonad was documented using the vital dyes acridine orange and ethidium bromide. Arch. Insect Biochem. Physiol. 36:95–114, 1997. © 1997 Wiley-Liss, Inc.

Low Temperature Effect on Viability of Diadegma insulare (Hymenoptera: Ichneumonidae) Pupae and Effect of This Parasitoid on Feeding Rate of Diamondback Moth Larvae (Lepidoptera: Plutellidae)

Laboratory tests with Diadegma insulare (Cresson), a parasitoid of the diamondback moth (Plutella xylostella (L.)), were conducted to evaluate the viability of cocoons stored at 4°C for varying lengths of time and also the effects of parasitism by D. insulare on the feeding rate of diamondback moth larvae. The percentage of adults emerging from cocoons declined steadily with time spent in storage at 4°C and was predictable (r2=0.9603; Y=98.3-2.06X; P<0.01). No emergence of D. insulare was observed after 49 d in storage. There was a significant difference in feeding rate of parasitized larvae and non-parasitized larvae. Consumption of collard foliage by parasitized larvae (86.25% of them were parasitized) the first day after stinging was about the same as for non-parasitized larvae; but the parasitized larvae then became sluggish and fed very little on days 2-5 when the experiment was terminated.

The effects of parasitism by Glyptapanteles liparidis (Braconidae: Hymenoptera) on the hemolymph and total body composition of gypsy moth larvae (Lymantria dispar, Lymantriidae: Lepldoptera).

The hemolymph and total body tissue composition (carbohydrates, lipids, proteins, and free amino acids) of gypsy moth larvae (Lymantria dispar, Lymantriidae:Lepidoptera) were analyzed and compared with the composition of larvae infected by their main parasitoid, Glyptapanteles liparidis (Braconidae:Hymenoptera). In the body tissue the concentrations of total lipids and total proteins decreased, whereas the glycogen concentration in the total body tissue was significantly elevated and exceeded the concentration of the nonparasitized larvae by a factor of approximately 3. Trehalose, glucose, and sorbitol were found in the hemolymph of both parasitized and nonparasitized larvae, but levels of all three carbohydrates were significantly reduced in parasitized individuals. The total lipid concentration was not altered due to parasitism. In the hemolymph the concentrations of total proteins nearly doubled in parasitized larvae, whereas the total free amino acid concentration was reduced; however, the levels of some single amino acids were reduced and those of others were elevated in the parasitized larvae. The polypeptide profile of the host hemolymph provided one parasitism-specific polypeptide at approximately 80 kDa. The results demonstrate that parasitization of gypsy moths by G. liparidis leads to significant alterations in levels of nearly all substance classes within the host's hemolymph and tissue.

Mechanism of parasitism-induced elevation of dopamine levels in host insect larvae

Parasitization with the parasitoid wasp, Cotesia kariyai, causes elevation of dopamine (DA) levels in the haemolymph and nerve cord of the last larval instar of the armyworm, Pseudaletia separata. Injection of DA or DA aganist [(−)-Quinpirole] during the early stages of the last larval instar retards normal growth and delays pupation. This suggests the possibility that DA plays an important role in the retardation of larval development by parasitism. The present study demonstrated that the DA level in the integument was also increased through the elevation of dopa decarboxylase (DDC) activity during parasitism. The total DA content in the integument was calculated to be approximately 50 times greater than in the haemolymph of parasitized Day 1 last instar larvae, thereby suggesting that parasitism-induced elevation of haemolymphal DA could be from DA in the integument. This hypothesis was partly confirmed by evidence that radiolabelled DA in the integument was secreted into in vitro incubation medium. Furthermore, DA elevation due to an increase in DDC activity in the integument was also observed in nonparasitized larvae in their last instar after injection of an insect biogenic peptide, growth-blocking peptide (GBP), whose concentration is elevated by parasitism. Therefore it is proposed that the plasma GBP fiter elevated by parasitism causes the rise in integumental DDC activity and this induces an increase in the DA level in the integument. This integument secretes DA into the haemolymph, raising DA levels in the haemolymph of parasitized larvae.

Parasitism by Microplitis demolitor induces alterations in the juvenile hormone titers and juvenile hormone esterase activity of its host, Pseudoplusia includens

Microplitis demolitor is a polydnavirus-carrying parasitoid that attacks the larval stage of Pseudoplusia includens and other noctuids. Parasitism or injection of wasp components like polydnavirus and teratocytes has a juvenilizing effect on P. includens development. Here we measured hemolymph juvenile hormone (JH) titers and juvenile hormone esterase activity in P. includens larvae after parasitism or injection of wasp components. Results were compared to nonparasitized larvae. During the fifth stadium, JH titers of nonparasitized larvae fluctuated between 0.08 and 0.50 ng/ml, whereas titers in parasitized larvae never fell below 2 ng/ml. P. includens larvae injected with calyx fluid plus venom or calyx fluid plus venom and teratocytes exhibited JH titers intermediate between parasitized and nonparasitized larvae. Nonparasitized larvae exhibited two peaks of JH esterase activity. The first occurred between 12 and 48 h in association with cessation of larval feeding, while the second occurred immediately before pupation. In contrast, levels of JH metabolism remained at low levels in parasitized larvae and larvae injected with calyx fluid plus venom or calyx fluid, venom and teratocytes. Only trace amounts of JH were detected in in vitro assays of teratocytes or M. demolitor larvae suggesting neither released JH into the hemolymph of parasitized larvae. Although calyx fluid plus venom and teratocytes did not elevate JH titers to the levels measured in parasitized larvae, their effects on formation of larval-pupal intermediates by P. includens were similar to those generated by exogenous application of methoprene or JH.

Effect of Parasitization by Cotesia kariyai (Hymenoptera: Braconidae) on Susceptibility of Pseudaletia separata (Lepidoptera: Noctuidae) Larvae to an Entomopoxvirus

Susceptibility of Pseudaletia separata larvae parasitized by Cotesia kariyai to an entomopoxvirus of P. separata (PsEPV) was compared to that of nonparasitized larvae under laboratory condition. Prasitized larvae were more susceptible than nonparasitized larvae when fourth-instar larvae were perorally inoculated with Ps EPV and then parasitized at 3 days later. The LD50 values for parasitized and nonparasitized larvae were 6.9×103 and 2.4×104 spheroids/ insect respectively and were significantly different as indicated by lack of overlap of their 95% fiducial limits. The presence of the wasp greatly enhanced the susceptibility of Pseudaletia separata larvae to the virus infection.

Precocious induction of metamorphosis inSpodoptera littoralis (noctuidae) by the parasitic waspChelonus inanitus (braconidae): Identification of the parasitoid larva as the key regulatory element and the host corpora allata as the main targets

The solitary egg-larval parasitoids of the genus Chelonus induce in their lepidopterous hosts precocious onset of metamorphosis followed by developmental arrest in the precocious prepupa. The parasitoid larva emerges as a freshly moulted 3rd instar larva from the precocious prepupa. In the Chelonus inanitus-Spodoptera littoralis system, we recently showed by using both direct and indirect approaches that polydnavirus and venom induce developmental arrest in the prepupa but do not induce precocious onset of metamorphosis [Soller and Lanzrein, J. Insect Physiol., in press]. Here we show results of parasitoid removal experiments and of experiments where parasitoid larvae were implanted into polydnavirus and venom containing larvae (in the absence of these components implanted parasitoids were encapsulated within a few hours). All our data indicate that the presence of a late first instar parasitoid larva is a prerequisite for precocious induction of metamorphosis in the 5th stadium. Under experimental conditions, the parasitoid larva often also induced the formation of miniature 6th instar larvae, which have a smaller head capsule width and reach a lower weight than nonparasitized larvae, suggesting that modification of threshold size for metamorphosis by the parasitoid larva is a gradual effect. Measurements of juvenile hormone biosynthesis by the corpora allata in vitro revealed that in parasitized larvae the corpora allata become precociously inactivated at the moult to the 5th stadium. Implantation of late first instar parasitoid larvae into 4th instar polydnavirus and venom containing larvae led to inactivation of the ricipient's corpora allata. These results suggest that in this parasitoid-host system factor(s) from the late first instar parasitoid larva inactivate the host's corpora allata which in turn leads to a precocious onset of metamorphosis. © 1996 Wiley-Liss, Inc.

Down-regulation of juvenile hormone esterase and arylphorin production inHeliothis virescens larvae parasitized byMicroplitis croceipes

Previous studies have shown that juvenile hormone esterase (JHE) activity and arylphorin titer were dramatically reduced in the hemolymph of Heliothis virescens larvae parasitized by Microplitis croceipes. Similar changes were observed in calyx fluid-injected or tetratocyte-injected larvae. In this study, we showed that the mRNA levels of both JHE and arylphorin transcripts were reduced significantly in fat body of parasitized larvae compared to those in nonparasitized larvae. The reduction of mRNA level appears to be fat body-specific, because no reduction of arylphorin mRNA level was observed in testes of the same parasitized larvae. Significantly, the mRNA levels of JHE and arylphorin in calyx fluid-injected or teratocyte-injected larvae were not different from those in control larvae, although the activity of JHE and the protein level of arylphorin were substantially lower in the treated larvae. Thus, our data suggest that both suppressed transcription of JHE- and arylphorin-encoding genes and post-transcriptional/translational modification(s) of JHE and arylphorin mRNAs or proteins may be involved in the reduction of JHE and arylphorin levels in parasitized H. virescens larvae. Calyx fluid and teratocytes may negatively regulate JHE and arylphorin levels via a transcriptional/translational mechanism. The factor(s) responsible for the reduction of JHE and arylphorin transcript levels by parasitization appears to be distinct from those induced by injection of calyx fluid or teratocytes. © 1996 Wiley-Liss, Inc.

Molecular cloning and characterization of cDNA for insect biogenic peptide, growth-blocking peptide

Growth-blocking peptide (GBP) is an insect biogenic peptide that prevents the onset of metamorphosis from larva to pupa. A cDNA coding for GBP is described. Mixed oligonucleotides derived from a GBP peptide sequence were used to generate amplified DNA by the polymerase chain reaction (PCR). Based on the sequence of the amplified DNA, a 41 bases oligonucleotide was designed for screening a cDNA library which was constructed from the armyworm Pseudaletia separata larvae parasitized with the parasitic wasp Cotesia kariyai. The cloned cDNA for GBP was 809 base pairs in length. An open reading frame of 429 base pairs encodes a pre-pro-peptide of 143 amino acid residues in which GBP is localized at the C-terminal region, and other three peptides including a putative signal peptide and appropriate processing sites for endoproteolytic cleavage precede the GBP sequence. Northern blot analyses demonstrate the presence of a 800-base mRNA transcript in fat body and 2.5-kilobase transcript in brain and nerve cord, suggesting the possibility that the transcription of GBP gene is regulated in a tissue-dependent manner. This interpretation was supported by isolating a GBP cDNA fragment from cDNA pool of brain-nerve cords. GBP mRNA is constantly expressed in both parasitized and non-parasitized last instar larvae and there is no difference in the levels of the mRNA between both larvae, thus indicating that parasitism may effect on translational or posttranslational level to elevate plasma GBP concentration.

Degradation of juvenile hormone III in vitro by non-parasitized and parasitized Spodoptera exigua (Noctuidae) and by the endoparasitoid Chelonus inanitus (Braconidae)

Parasitism of Spodoptera exigua by the solitary egg-larval parasitoid Chelonus inanitus induces precocious onset of metamorphosis, i.e. parasitized larvae construct a pupal cell and become prepupae in the fifth larval instar while non-parasitized larvae do so in the sixth larval instar. The parasitoid larva emerges as a freshly molted third instar larva from the precocious prepupa. A comparison of the metabolism of racemic 3H-JH III by homogenates of non-parasitized and parasitized larvae at the stage of pupal cell formation revealed that very similar amounts of juvenile hormone III diol, juvenile hormone III acid and juvenile hormone III acid-diol were formed. This shows that the changes in juvenile hormone degradation associated with metamorphosis, which occur in the sixth stadium in non-parasitized larvae, are precociously induced in parasitized larvae in the fifth stadium. In homogenates of parasitoid larvae, juvenile hormone III metabolizing activity was very low during the internal phase of development (first to second instar larvae) and almost excusively juvenile hormone III diol was formed. After emergence from the host, i.e. in third instar larvae, juvenile hormone III metabolic activity increased and esterases became active. In comparison to the host, specific juvenile hormone III degrading activities were always much lower in homogenates of parasitoid larvae than of host larvae. With homogenates of S. exigua eggs at a stage 2 3 of embryonic development and shortly before hatching, the pattern of juvenile hormone III metabolism was almost identical whether parasitized or not, and the major metabolite being formed was juvenile hormone III diol. This shows that parasitism by C. inanitus does not influence juvenile hormone metabolic activities at these stages.

An Abundantly Expressed Hemolymph Glycoprotein Isolated from Newly Parasitized Manduca sexta Larvae Is a Polydnavirus Gene Product

Cotesia congregata polydnavirus (CcPDV) is essential for successful parasitism of Manduca sexta larvae by the braconid wasp C. congregata. CcPDV virions are present in large numbers in the oviducts of C. congregata and injected with eggs into the hemocoel of M. sexta larvae during parasitization. Injection of sucrose density purified virions into nonparasitized larvae causes several of the parasitism-induced alterations in the physiology of host larvae that occur as a result of natural parasitism, including the synthesis of novel hemolymph proteins and abrogation of the host's immune response against the developing parasites. One of these proteins, early-expressed protein 1 (EP1), is a 190-kDa molecule which constitutes up to 5% of the total hemolymph protein by 24 hr following oviposition by the wasp. Using N-terminal sequence data for EP1 to construct primers for use in the polymerase chain reaction, we amplified and cloned a cDNA corresponding to the gene encoding EP1. This cDNA hybridized to DNA of the CcPDV genome, but not to DNA isolated from M. sexta larvae, suggesting that EP1 is a CcPDV gene product. A cDNA clone was isolated from an expression library generated from RNA extracted from newly parasitized M. sexta larvae. Sequence analysis of the cDNA clone revealed the presence of an open reading frame of 819 bp encoding a protein of 30.7 kDa. In vitro transcription/translation of the cDNA clone produced a protein of approximately 31 kDa, which was immunoprecipitated by EP1-specific polyclonal antiserum generated against purified deglycosylated EP1. EP1-like sequences also were amplified from male wasp genomic DNA, suggestive of integration of EP1-like sequences in the genome. This report constitutes the first evidence that a specific protein isolated from a parasitized host insect is a wasp polydnavirus gene product.

Parasitism of Pickleworm and Melonworm (Lepidoptera: Pyralidae) by Cardiochiles diaphaniae (Hymenoptera: Braconidae)

Pickleworm, Diaphania nitidalis (Stoll), and melonworm, Diaphania hyalinata (L.), larvae were susceptible to parasitism by Cardiochiles diaphaniae Marsh. Melonworm larvae were more susceptible to parasitism than pickleworm, but both exhibited reduced weight gain and head capsule growth in later stadia compared with nonparasitized conspecifics and then perished following initiation of wandering before pupal molt. Melonworm larvae parasitized when in advanced stadia achieved a greater final weight than larvae parasitized when young. Parasitoid larval development time was longest following parasitization of young melonworm, but not significantly different in melonworm larvae parasitized in middle or later instars. Compared with nonparasitized melonworm larvae, the prepupal wandering stage was abbreviated in melonworm larvae parasitized when young, and extended in melonworm larvae parasitized in later instars. Overall, parasitoid development time was significantly longer in pickleworm hosts than melonworm. Adult parasitoid weight increased with host age at the time of parasitization when melonworm served as host and was highest in middle-instar larvae when pickle worm served as host. C. diaphaniae developing in pickleworm larvae weighed significantly more than those developing in melonworm larvae.

Purification and characterization of an early-expressed polydnavirus-induced protein from the hemolymph of Manduca sexta larvae parasitized by Cotesia congregata

Parasitism of Manduca sexta by Cotesia congregata causes major qualitative and quantitative changes in the host's hemolymph proteins, with several novel parasitism-specific proteins being expressed in newly infected larvae. This report describes evidence for the production of several early-induced proteins (EP's) all of which have subunit molecular weights in the range of 30,000–40,000 and appear in the hemolymph of M. sexta within a few hours following parasitization. Injection of purified polydnavirus into non-parasitized larvae induced production of several EP proteins, suggesting that onset of their synthesis is virally mediated. Following analysis of hemolymph proteins on 2-dimensional gels, N-terminal sequence data were obtained for three EP proteins and one was selected for further study, which was designated EP1. This protein was purified to homogeneity using gel filtration and chromatofocusing; treatment of native purified protein with glycosidases showed that EP1 was heavily glycosylated with polysaccharides of the high-mannose type and the denatured, deglycosylated protein had a subunit molecular weight of c. 33,000. The native molecular weight of EP1 was estimated by gel filtration and native gradient polyacrylamide gel electrophoresis (PAGE) c. 190,000. Western blots of hemolymph from parasitized larvae showed that the EP1 antisera detected proteins only in parasitized larvae; the antibodies showed no cross-reactivity with other proteins present in the hemolymph of non-parasitized larvae. The Western blots indicated the early protein(s) were immunologically detectable in the hemolymph from 2 to 144 h following parasitization. Slot blots using a labeled internal restriction fragment from an early protein-specific cDNA (believed to be EP1 cDNA) as probe showed that transcripts from fat body of parasitized larvae were detectable from 0.5 to 96 h post-oviposition, suggesting that synthesis of transcripts rapidly occurs. At 24 h post-oviposition, these transcripts were detected primarily in fat body and hemocytes; lower amounts were also detected in Malpighian tubules and midgut tissues.

Alteration of ecdysone metabolism in Heliothis virescens (F.) (Lepidoptera: Noctuidae) larvae induced by Cardiochiles nigriceps Viereck (Hymenoptera: Braconidae) teratocytes

The haemolymph titres of ecdysteroids in last instar Heliothis virescens (F.) larvae parasitized by Cardiochiles nigriceps Viereck, or in larvae injected with teratocytes obtained from this parasitoid, were determined and compared to those of nonparasitized larvae. Ecdysteroids were extracted, purified by high performance liquid chromatography (HPLC), and the titres assessed by radioimmunoassay. The total ecdysteroid titre of parasitized day 5 of 5th instar larvae was consistently lower than in nonparasitized larvae of the same age. The main ecdysteroid inactivation products were both polar and C-26 hydroxylated compounds, which were detected as soon as the 20-hydroxyecdysone appeared to be produced. The subsequent increase of the total ecdysteroid titre registered on day 7 of 5th instar parasitized larvae was almost completely due to polar compounds, whereas 20-hydroxyecdysone remained at low levels. In contrast, the total ecdysteroid titre of nonparasitized H. virescens larvae was higher and inactive metabolites increased in concentration only the day before pupation, after the typical 20-hydroxyecdysone burst. When C. nigriceps teratocytes, obtained from embryos hatched in vitro on a semi-defined medium, were injected into day 1 of nonparasitized host 5th instar larvae (new-slender stage), the total ecdysteroid titres on day 5 and 6 of these larvae were much higher than in nonparasitized controls. However, this high ecdysteroid titre was associated with low levels of 20-hydroxyecdysone and with a high titre of ecdysone and other unidentified ecdysteroids. The 20-hydroxyecdysone was the most abundant ecdysteroid released after enzymatic digestion of the polar ecdysteroids isolated from the haemolymph of parasitized day 7 of 5th instar larvae. In vitro incubation of teratocytes with radiolabelled 20-hydroxyecdysone resulted in the recovery of 34% of the total radioactivity in the HPLC fractions corresponding to polar metabolites. In contrast, only 4.5% of the total radioactivity was recovered in the same polar fractions when ecdysone was co-incubated with teratocytes. These results, along with those presented above, suggest that the inhibition of host pupation observed in parasitized H. virescens larvae is due in part to 20-hydroxyecdysone inactivation through the formation of polar metabolites and C. nigriceps teratocytes seem to play an important role in this biochemical transformation.

Premature production of late larval storage proteins in larvae of trichoplusia ni parasitized by euplectrus comstockii

AbstractInvestigations were conducted to determine the titer of storage proteins in larvae of the cabbage looper, Trichoplusia ni (Hübner), that were parasitized by the ectoparasitoid Euplectrus comstockii Howard (Hymenoptera: Eulophidae). A gradual increase was noted in the titer of the storage proteins present in the hemolymph of parasitized third and fourth instar larvae and in the hemolymph of isolated thoracic and abdominal tissues of fourth instar larvae. The final amount present in parasitized third and fourth instar larvae was similar to that found in nonparasitized fifth instar larvae. The stimulation of storage proteins in envenomed larvae demonstrates the ability (competence) of early larval stages to produce a gene product that normally occurs in the last larval stadium of the lepidopteran larval host. The gene expression necessary for storage protein production in isolated tissues may be altered by mechanisms separate from inherent developmental processes and the intact endocrine system. © 1994 Wiley‐Liss, Inc.This article is a US Government work and, as such, is in the public domain in the United States of America.

Pesticides Affect Immature Stages of Diadegma insulare(Hymenoptera: Ichneumonidae) and Its Host, the Diamondback Moth (Lepidoptera: Plutellidae)

Effects of pesticides on immature stages of Diadegma insulare (Cresson) and its host, the diamondback moth, Plutella xylostella (L.), were studied in the laboratory. Diamondback moth larvae parasitized by D. insulare were significantly less sensitive to ingested pesticides than were nonparasitized larvae 48 h after treatment. However, they were equally sensitive to pesticides through contact. Bacillus thuringiensis var. kurstaki (Berliner) and azinphosmethyl were significantly more toxic to parasitized larvae than permethrin, methomyl, and chlorothalonil if ingested. D. insulare adults emerging from pupae that formed from larvae surviving pesticide treatments tended to have more females than those in the control group. Except for B. thuringiensis and chlorothalonil treatments, percentage adults emerging from pesticide-treated pupae was significantly lower for D. insulare than for diamondback moth in either direct-dip or spraying bioassays. Lower pesticide sensitivity in immature stages of D. insulare may be important for effective integrated diamondback moth management.

Effects of parasitism by the braconid wasp Cotesia congregata on host hemolymph proteins of the tobacco hornworm, Manduca sexta

Parasitism by the braconid wasp Cotesia congregata causes major alterations in the hemolymph proteins of host tobacco hornworm larvae. Earlier studies showed that the total amount of hemolymph protein is reduced during parasitism, beginning almost immediately after the host is parasitized. Simultaneously, parasitism induces synthesis of large amounts of novel proteins that appear in the blood as early as 1–2 h post-parasitization. The present report confirms earlier studies describing the presence of novel proteins in last instar hosts, and also characterizes the effects of parasitism in altering the titers of several endogenous host hemolymph proteins normally produced by the fat body and other tissues. Analysis of hemolymph plasma using SDS-PAGE and densitometry, as well as immunodiffusion assays, showed that in terminal stage fifth instar host larvae, the titers of serpins and arylphorin were dramatically reduced relative to the levels of these proteins detected in nonparasitized gate II fifth instar larvae of the same age. The relative differences between parasitized and nonparasitized larvae increased with time following ecdysis to the fifth instar, so that the day 4 nonparasitized larvae had arylphorin titers of c. 30 mg/ml, whereas parasitized day 4 larvae with newly emerged wasps had only one sixth that amount of storage protein circulating in the hemolymph. Similarly, in nonparasitized larvae the hemolymph serpin concentration increased from c. 200 μg/ml (on day 0) to > 600 μg/ml (on day 4) in prewandering gate II larvae, but in parasitized larvae the hemolymph serpin concentration was maintained in the range of 100–200 μg per ml hemolymph until the pharate third instar parasites emerged from the host larva on day 4. In contrast, the level of hemolymph lipophorin was unaffected by parasitism, and lipophorin increased from c. 1.3 to > 3 mg/ml during the time interval between days 0 and 4 in both nonparasitized and parasitized larvae. Hemolymph titers of insecticyanin also were not significantly different in parasitized vs nonparasitized larvae, and in both types of larvae the concentration of this pigment decreased by c. 50% during the same time interval when lipophorin was increased significantly. Instead of causing a generalized inhibition of host hemolymph protein synthesis, parasitism causes a complex array of changes in the hemolymph protein profile of Manduca sexta, possibly via the mediation of hormonal modulators of host protein synthesis, or transcriptional or translational regulation of host gene expression by factors associated with the polydnavirus or molecules secreted by the parasites. Another explanation might be that the turnover rates for serpins and arylphorins differ in parasitized vs unparasitized larvae. These two interpretations could not be distinguished on the basis of the data reported here.

Expression of a parasitism‐specific protein in lepidopteran hosts of Chelonus sp.

The hemolymph of each noctuid species successfully parasitized by Chelonus near curvimaculatus possessed a parasitism-specific protein (PSP) previously identified in host T. ni (Insect Biochem. 19:445;21:845). Expression of PSP occurred in a stage-specific manner in the stadium during which the host undergoes precocious metamorphosis. The appearance of the protein was not due to nutritional stress associated with parasitism of hosts, since starved nonparasitized larvae did not produce the protein, or to low juvenile hormone titers occurring in precociously metamorphosing hosts, but rather was dependent on the presence of the endoparasite larva. Results of an in vivo incorporation experiments with [35S]-methionine showed that synthesis and subsequent appearance of the protein in the hemolymph of parasitized hosts was abrogated by prior surgical removal of endoparasite. Immunoprecipitation analysis of proteins from C. near curvimaculatus larvae cultured in vitro using antibodies specific to PSP indicated that the source of the protein was the endoparasite. Synthesis of PSP by the endoparasitic larvae with its subsequent secretion into the hemocoel of hosts was specific to the advanced stages of parasite development prior to its regression from the host.

CONSERVATION OF SPRUCE BUDWORM PARASITOIDS FOLLOWING APPLICATION OF BACILLUS THURINGIENSIS VAR. KURSTAKI BERLINER

AbstractField trials in northern Ontario in 1990 and 1991 confirmed that survival of spruce budworm larvae [Choristoneurafumiferana (Clemens)] (Lepidoptera: Tortricidae) parasitized by Apanteles fumiferanae Viereck (Hymenoptera: Braconidae) was greater than survival of nonparasitized budworm larvae following application of Bacillus thuringiensis var. kurstaki Berliner (Bt). Apparent parasitism by A. fumiferanae increased slightly when application of Bt was early (instar III or IV) but increased significantly when applications were later in the season (instars IV–VI). Delaying the timing of the spray conserved parasitoids while providing significant reductions in budworm densities and not compromising the primary objective of Bt application, i.e. the protection of current year foliage. The potential for long-term management of spruce budworm through the strategic integration of natural and imposed mortalities is discussed.