Host Spodoptera Littoralis Research Articles

Improved reproduction of Chelonus inanitus on Ephestia kuehniella highlights this factitious host as an alternative to the natural host Spodoptera littoralis

Chelonus inanitus L. (Hymenoptera: Braconidae) is an egg-larval parasitoid of Lepidoptera, with the potential for biological control of noctuids. The lack of information merits research on its reproductive and developmental biology. We studied the reproduction of mated parasitoid pairs during female lifetime and the development of the progeny on two hosts, the factitious Ephestia kuehniella Zeller (Lepidoptera: Pyralidae) and the natural Spodoptera littoralis Boisduval (Lepidoptera: Noctuidae). The longevity of pairs subjected or not to parasitism was compared. Development of the progeny after parasitization and growth rates were studied. Reproduction decreased the longevity of both sexes in every host. The similar survival patterns proved that sex was not a factor influencing longevity. Females parasitized immediately after emergence and offspring production peaked at 2–2.5 days. Low parasitization rates (3–12%) were generally observed. Reproduction on S. littoralis was fourfold less efficient, probably due to failure of immature stages leading to poor progeny. Sex ratio of the progeny was mostly male-biased. Males developed faster in both hosts. Development was 20 days shorter in the natural host. Strikingly, the parasitoid performed successfully better on the factitious host. Inbreeding, poor understanding of host preference, and inadequate rearing and handling practices are discussed as possible causes of the worse parasitoid performance on S. littoralis.

Midgut microbiota and host immunocompetence underlie Bacillus thuringiensis killing mechanism

Bacillus thuringiensis is a widely used bacterial entomopathogen producing insecticidal toxins, some of which are expressed in insect-resistant transgenic crops. Surprisingly, the killing mechanism of B. thuringiensis remains controversial. In particular, the importance of the septicemia induced by the host midgut microbiota is still debated as a result of the lack of experimental evidence obtained without drastic manipulation of the midgut and its content. Here this key issue is addressed by RNAi-mediated silencing of an immune gene in a lepidopteran host Spodoptera littoralis, leaving the midgut microbiota unaltered. The resulting cellular immunosuppression was characterized by a reduced nodulation response, which was associated with a significant enhancement of host larvae mortality triggered by B. thuringiensis and a Cry toxin. This was determined by an uncontrolled proliferation of midgut bacteria, after entering the body cavity through toxin-induced epithelial lesions. Consequently, the hemolymphatic microbiota dramatically changed upon treatment with Cry1Ca toxin, showing a remarkable predominance of Serratia and Clostridium species, which switched from asymptomatic gut symbionts to hemocoelic pathogens. These experimental results demonstrate the important contribution of host enteric flora in B. thuringiensis-killing activity and provide a sound foundation for developing new insect control strategies aimed at enhancing the impact of biocontrol agents by reducing the immunocompetence of the host.

Toxicity and kinetics of spinosad in different developmental stages of the endoparasitoid Hyposoter didymator (Hymenoptera: Ichneumonidae) and its host Spodoptera littoralis larvae (Lepidoptera: Noctuidae)

The toxicity of spinosad was evaluated using the RaPID Assay® Spinosad immunosorbent assay in different developmental stages of the parasitoid, Hyposoter didymator, and in its host, fourth-instar larvae of the cotton leafworm Spodoptera littoralis. Spinosad was applied directly to pupae and adults of H. didymator (ingestion or topical application) or to the immature stages of the parasitoid via the host larvae. Low amounts of spinosad were recovered from S. littoralis host larvae after topical treatment, and the compound was mainly retained in the hemolymph. Amounts of spinosad detected in third-instar larvae of H. didymator, pulled out from the hemolymph of parasitized S. littoralis larvae, were 85 pg (3.57 ng a.i./g body weight) in dead larvae, and 82 pg (3.42 ng a.i./g body weight) in alive individuals. After topical treatment of H. didymator cocoons, most of the compound was retained in the silken cocoon, preventing contamination of the pupa. Also in the parasitoid adults, relatively low amounts of spinosad were accumulated in the body overall, but half of all the insecticide recovered was found in the ovaries. The kinetic results obtained help to better understand the toxicity of spinosad in the complex S. littoralis–H. didymator, and to ascertain the compatibility between spinosad and the parasitoid for optimizing the control of lepidopteran pests.

The influence of two endoparasitic wasps, Hyposoter didymator and Chelonus inanitus, on the growth and food consumption of their host larva Spodoptera littoralis

The influence of parasitism by Hyposoter didymator (Thunberg; Hymenoptera: Ichneumonidae) and Chelonus inanitus (Linnaeus) (Hymenoptera: Braconidae) on the growth and food consumption of their host Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae) was studied in the laboratory. Parasitised larvae consumed significantly less artificial diet than unparasitised ones. Egg parasitisation by C. inanitus affected host larval consumption from the second day after emergence and it was significantly different from that of unparasitised ones. H. didymator, however, started to reduce larval consumption 4 days after parasitisation on the third instar host larvae. The overall reduction achieved by the larval endoparasitoid H. didymator is higher than that caused by the egg-larval endoparasitoid C. inanitus. The final body weight of a parasitised host larva by H. didymator and C. inanitus was only 6.7 and 13.0% of the maximum weight of an unparasitised sixth instar larva respectively. Moreover, parasitised larvae never reached the last instar. Results indicated that parasitised larvae might cause considerable less damage to the host plant than unparasitised ones.

The calyx fluid of Microplitis rufiventris parasitoid and growth of its host Spodoptera littoralis larvae

The morphology of the female reproductive system of pupal and adult stages of Microplitis rufiventris and the ultrastructure of the ovaries are described and illustrated. Two morphologically distinct types of particles of nuclear origin, i.e., polydnavirus (PDV) and a virus-like filamentous particle (VLFP) were detected in the ovarian calyx fluid of the female wasp. It is likely that these particles are injected into the host during oviposition. PDV initiated replication in the calyx of mid-aged pupae and in pharate adults and were present throughout adult life. VLFP were only seen in the calyx fluid of newly emerged adults, and therefore observed after the PDV. Feulgen and methyl-green pyronin staining revealed the presence of DNA in both types of particles. The effects of injection of Spodoptera littoralis larvae with a combination of parasitoid viruses and venom of M. rufiventris females (CxFV) were investigated and the results were compared with two control groups, i.e., larvae injected with Pringle's saline (PS) and naturally parasitized larvae. CxFV-larvae showed significant declines ( P < 0.05 ) in food consumption, weight of ejected faeces and weight gain when compared with PS-larvae. However, naturally parasitized larvae (parasitoid egg+CxFV+ovarian protein) displayed a high significance score ( P < 0.01 ) in comparison with those of PS-larvae. The approximate digestibility (AD) values of S. littoralis larvae were positively affected as early as day 2 post-treatment by either injection of CxFV or parasitization. However, a reduction in AD was observed in both PS- and CxFV-larvae on day 3–7 in comparison with naturally parasitized larvae. Other indices of food utilization were unchanged in CxFV-larvae when compared to saline treated or parasitized controls.

Changes in the haemolymph proteome of Spodoptera littoralis induced by the parasitoid Chelonus inanitus or its polydnavirus and physiological implications

The egg–larval parasitoid Chelonus inanitus induces in its host Spodoptera littoralis two major developmental effects, namely a precocious onset of metamorphosis followed by a developmental arrest in the prepupal stage. Along with each egg, the wasp injects polydnavirus and venom into the host egg. The polydnavirus has been shown to play a major role in inducing the developmental arrest while the parasitoid larva is instrumental in inducing the precocious onset of metamorphosis. Here we report that experimental dilution of haemolymph of polydnavirus-containing larvae can partially prevent the developmental arrest while injection of native, but not of heat-treated, haemolymph or plasma from polydnavirus-containing larvae into nonparasitized larvae could induce developmental arrest in 14–15% of the larvae. This illustrates that heat-labile factors present in haemolymph play a role in causing developmental arrest. Injection of parasitoid medium increased the proportion of larvae entering metamorphosis precociously while injection of antibodies against a parasitoid-released protein had the opposite effect; this indicates that this protein and possibly other parasitoid-released substances are involved in inducing the precocious onset of metamorphosis. Analysis of the plasma proteome of nonparasitized, parasitized and polydnavirus-containing larvae revealed that the developmental effects are associated with only minor differences: eleven low abundant viral or virus-induced proteins and five parasitoid-released proteins were seen at specific stages of the host.

Fate of polydnavirus DNA of the egg–larval parasitoid Chelonus inanitus in the host Spodoptera littoralis

In situ hybridizations show that 5 min after parasitization, polydnavirus DNA is in close vicinity of the parasitoid egg, but 5 h later also in the yolk and partially in the host embryo. Fifteen hours after parasitization, the viral DNA is seen all over the host embryo and hardly in the yolk. The tissue distribution of the viral DNA was analysed and quantified by dot blots in the fifth instar parasitized larvae. On a per host basis, haemocytes and fat body contained the highest amount of viral DNA, while nervous tissue, intestinal tract and carcass contained less. Of the three viral segments tested, all were found in all tissues. Relative to the quantity of host DNA, viral DNA was most abundant in haemocytes, about five times less abundant in fat body and nervous tissue and about 25 times less abundant in intestinal tract. The total quantity of viral DNA per host was 444±145 pg which is similar to the quantity injected by the wasp; thus, the viral DNA persists throughout parasitization. The parasitoid larva contains 820±80 pg viral DNA integrated in the genome. This illustrates that the dose of viral DNA injected in virions represents approximately one third of the total viral genomic information present in a host at a late stage of parasitism.

Characterization of two genes of the polydnavirus of Chelonus inanitus and their stage-specific expression in the host Spodoptera littoralis.

Chelonus inanitus (Braconidae, Hymenoptera) is a solitary egg-larval parasitoid of Spodoptera littoralis. Along with the egg the female wasp injects polydnaviruses, which are prerequisites for successful parasitoid development. The polydnavirus genome is segmented and consists of double-stranded circular DNA. Proviral DNA is integrated in the wasp's genome; virus replication is restricted to the wasp's ovary and does not occur in the parasitized host. The polydnavirus of C. inanitus (CiV) protects the parasitoid larva from encapsulation by the host's immune system and causes a developmental arrest of the host in the prepupal stage. Here we report on the first two cloned CiV genes, which are named CiV14g1 and CiV14g2 because of their localization on segment CiV14. The cDNA of CiV14g1 has a size of 2036 bp; the gene contains seven exons interrupted by six introns of similar size and encodes a putative polypeptide of 548 amino acids. The cDNA of CiV14g2 has a size of 618 bp; the gene consists of three exons and encodes a putative peptide of 77 amino acids. Transcript quantities of both genes are very low up to the penultimate larval instar of the host. In the last instar, at the stage of pupal cell formation, CiV14g1 expression increases about 5-fold and CiV14g2 expression about a 1000-fold. These are the first data to show strong upregulation of polydnavirus genes towards the end of parasitization. These two genes might be involved in the reduction of host ecdysteroids observed at this stage.

Teratocytes of microplitis rufiventris in superparasitized host larvae

The effects of superparasitism on the size and number of teratocytes (giant cells released by the serosa of braconid eggs after hatching) found in the host Spodoptera littoralis at the completion of development of the solitary endoparasitoid Microplitis rufiventris were assessed. Although the number of teratocytes increased, on average, with parasitoid load (1, 2 or 3 parasitoid eggs), this increase was lower than predicted, based on the number of parasitoids (i.e., is not proportional to parasitoid load). Similarly, the size of the teratocytes decreased as a function of parasitoid load. The total volume of the teratocytes (mm3) is reduced by 39.1 and 66.4% in hosts parasitized twice and three times, respectively. It is concluded that, in superparasitized hosts, some form of competition likely takes place among the teratocytes, or between these cells and the first‐hatched larva, possibly through the action of a chemical (released by the latter) that inhibits the growth of teratocytes from supernumerary parasitoids. In singly parasitized hosts 73.2, 17.8 and 9.0% of total teratocytes were found in hosts abdomen, head capsule and thoracic regions, respectively. In superparasitized hosts, the cells tended also to collect in the posterior region of the host but with fewer number of cells in the head capsule. It seems that the teratocytes possibly act as a biological regulating factor to adjust host's endocrine system for successful development of the parasitoid.

Overview of parasitism associated effects on host haemocytes in larval parasitoids and comparison with effects of the egg-larval parasitoid Chelonus inanitus on its host Spodoptera littoralis

In the first part we review the effects of larval endoparasitoids and their polydnavirus and venom on the immune system of their hosts. In all systems investigated, haemocyte spreading and encapsulation activity was reduced; in some cases effects on total (THC) or differential (DHC) haemocyte count as well as modification of haemocyte morphology and ultrastructure were also documented. In many cases polydnavirus (and venom) were shown to play a major role in abrogation of the host's immune reaction. In the second part we present the first investigation of effects of parasitism and polydnavirus/venom on the immune system of the host for an egg-larval parasitoid, Chelonus inanitus. We observed that in 4th and 5th instar larvae, i.e. 7 to 10 days after parasitization, neither haemocyte spreading and encapsulation activity, nor DHC, nor haemocyte ultrastructure were altered. After parasitization with X-ray irradiated wasps, which inject polydnavirus and venom and infertile eggs, there was no alteration of the above mentioned parameters. Nevertheless, parasitoid larvae implanted into 4th instar larvae which developed from eggs parasitized with X-ray irradiated wasps were not encapsulated, whereas co-injected latex beads were. These results show that parasitism by this egg-larval parasitoid does not generally suppress the host's immune system but that polydnavirus/venom injected at oviposition prevent, by, as yet unknown mechanisms, encapsulation of the parasitoid larva.

Polydnavirus and venom of the egg-larval parasitoid Chelonus inanitus (Braconidae) induce developmental arrest in the prepupa of its host Spodoptera littoralis (Noctuidae)

The egg-larval parasitoid Chelonus inanitus induces in its host Spodoptera littoralis precocious onset of metamorphosis in the penultimate stadium and developmental arrest in the precocious prepupa. At oviposition the wasp injects, along with the parasitoid egg, venom and calyx fluid that contains polydnaviruses. By using indirect and direct approaches we have investigated the role of polydnaviruses and venom in influencing host development. When parasitoid development was prevented by either X-ray irradiation of wasp females or heat-treatment of parasitized eggs, hosts developed normally up to the prepupal stage but then became developmentally arrested. Injection of calyx fluid and venom into eggs also led to developmental arrest in the prepupa, while injection of anti-polydnavirus antibodies into eggs before parasitization prevented successful parasitoid development and rescued hosts so that they pupated normally. These results show that polydnavirus and venom induce developmental arrest in the prepupa but do not induce precocious onset of metamorphosis. Venom alone was ineffective in causing developmental arrest, and calyx fluid alone was effective only at high doses; but together with 0.0048 wasp equivalents of venom, as little as 0.0005 wasp equivalents of calyx fluid per egg was sufficient to induce developmental arrest. SDS-gel electrophoretic analyses of purified polydnavirus and venom showed that both consist of a great number of polypeptides of various sizes, and Western blots revealed that the anti-polydnavirus antibodies crossreacted with several venom proteins.

Biology and morphology of the parasitoidChelonus inanitus(Braconidae, Hymenoptera) and effects on the development of its hostSpodoptera littoralis(Noctuidae, Lepidoptera)

Summary The solitary egg-larval parasitoids of the genus Chelonus have the unique effect of inducing in their lepidopterous hosts the precocious onset of metamorphosis and developmental arrest in the precocious prepupa. As a prerequisite to identifying the physiological and biochemical mechanisms underlying these developmental effects, we analyzed the morphology and various growth parameters in the course of development of Chelonus inanitus and studied the effect of this parasitoid on growth and development of its host Spodoptera littoralis. We show the first scanning electron microscopic study of the internal stages of an endoparasitic wasp from the embryo to the third instar. By including observations from microscopic and histological analyses we describe the different stages in the development of the anal vesicle and the formation of new segments and show that the anal vesicle has surface structures which were never described before. We also demonstrate that the head and mouthparts change entirely with...

Cloning of the genome of a densovirus and rescue of infectious virions from recombinant plasmid in the insect host Spodoptera littoralis

We have cloned an infectious genome of the Junonia coenia densonucleosis virus (JcDNV) into the bacterial plasmid pBR322. The viral genome could be rescued from the recombinant plasmid pBRJ by transfection of pBRJ DNA to sensitive Spodoptera littoralis larvae. pBRJ DNA produced a typical viral infection and a comparable percentage of larvae became infected following inoculation of equivalent amounts of purified virion DNA or cloned viral DNA. Virions extracted from transfected larvae were indistinguishable from wild-type (wt) virions with regard to their biophysical and biological properties. In particular, rescued virions were as infectious as wt virions and showed identical restriction profiles of their genome. In contrast, subcloning of JcDNV DNA deleted at both extremities of a sequence of ca 250 or ca 100 by resulted in the inability of the recombinant plasmids to initiate a viral infection. These data suggest that, as for vertebrate parvoviruses, the inverted terminal repeats display essential functions in the rescue process and replicative cycle of densoviruses. This is the first report of the molecular cloning of the infectious genome from an insect parvovirus, and more generally from an invertebrate virus. pBRJ should provide an efficient tool to further define the organization of the JcDNV genome and compare it to other parvoviruses.