Persistence and expression of Cotesia congregata polydnavirus in host larvae of the tobacco hornworm, Manduca sexta.

The gregarious braconid wasp Cotesia congregata parasitizes host larvae of Manduca sexta, and several other sphingid species. Parasitism induces host immunosuppression due to the disruptive action of the wasp's polydnavirus (PDV) on host blood cells. During the initial stages of parasitism, these cells undergo apoptosis followed by cell clumping, which clears the hemolymph of a large number of cells. In this study, the persistence and expression of Cotesia congregata PDV (CcPDV) were examined using Southern and Northern blots, respectively. Digoxygenin-labelled total polydnaviral DNA was used to probe genomic DNA isolated from fat body and brains of hosts with emerged wasps taken 6 days following egress of the parasitoids, and significant cross-hybridization between the host fat body genomic DNA with viral DNA was seen. Thus, the virus persists in the host for the duration of parasitism, even during the post-emergence period, and may even be integrated in the host caterpillar DNA. Viral gene expression was examined using Northern blots and probes to the Cotesia rubecula CrV1 homolog, and the CrV1-like mRNAs were expressed as early as 4 h post-parasitization for at least 72 h and faint hybrization is even seen at the time the wasps eclose. In contrast, in Pieris rapae larvae the CrV1 transcript is expressed only for a brief time, during which time hemocyte function is disrupted. The effect is transitory, and hemocytes regain their normal functions after the parasites emerge as first instars.The genome of CcPDV contains one copy of the CrV1-like homolog as shown on Southern blots of viral genomic DNA. In conjunction with our earlier studies of the PDV-encoded early protein 1, the current work suggests multiple viral transcripts are produced following parasitization of the host, and likely target host hemocytes to induce their apoptosis, thereby preventing encapsulation of the parasitoid's eggs. Whether viral DNAs are integrated in the host's genomic DNA remains to be proven, but our results provide preliminary evidence that viral DNAs are detected in the host's fat body cells examined at the time of wasp emergence and several days later.

Parasitoid-host endocrine relations: self-reliance or co-optation?

High titers of juvenile hormone (JH) maintain developmental arrest in Manduca sexta larvae parasitized by Cotesia congregata. Parasitized hosts exhibit up to 9.5 times greater amounts of total hemolymph JH (from 0.6+/-0.09 to 2.51+/-0.43ng/ml) compared to non-parasitized controls. Elevated titers are observed throughout the fifth instar, even beyond egression of the parasitoids on day 5. GC-MS analysis revealed that in hemolymph of unparasitized control larvae, JH I is the major homolog and levels of JH III are negligible; in parasitized individuals the amounts of JH I, II, and III rise, and JH III predominates. Neck ligation ensured separation of M. sexta's corpora allata from the posterior section, which contained most of the parasitoids in the infected insects. When the posterior region was sampled, JHs were not detected in the non-parasitzed larvae, but in those parasitized, JH III was found (1.98+/-0.29ng/ml, 24 h post-ligation). JH III was the only homolog produced and secreted by the parasitoid in in vitro culture. This is the first report stating that a parasitoid secretes JH III and may contribute, at least in part, to the circulating titer in the host hemocoel, concurrently promoting host production of JH I and II.

Co-infection of Manduca sexta larvae with polydnavirus from Cotesia congregata increases susceptibility to fatal infection by Autographa californica M Nucleopolyhedrovirus.

We investigated pathogenesis of Autographa californica M Nucleopolyhedrovirus in the semipermissive host, Manduca sexta, using a lacZ recombinant virus (AcMNPV-hsp70/lacZ) to track the temporal progression of infection. Results from time course studies monitoring infections initiated orally in fourth instars demonstrated that primary infection of midgut columnar cells began at 3 h post inoculation (hpi). We observed secondary infections in midgut-associated tracheae as early as 9 hpi, showing that the early events of pathogenesis in M. sexta are similar to those of permissive noctuid larvae. In M. sexta, however, unlike in permissive hosts, hemocytes rapidly surrounded infected tracheal cells and formed capsules. Subsequently, baculovirus infections failed to spread and ultimately were cleared, suggesting that a cellular immune response had been triggered. To assess the effects of immunosuppression on baculovirus-induced disease, we compared the outcome of infections in immunocompetent hosts with those that were immunocompromised either by parasitization with the braconid, Cotesia congregata, or by injection of the parasitoid's polydnavirus. During the first 9 days after inoculation, parasitized and polydnavirus-inoculated M. sexta larvae died more quickly and at higher levels than nonparasitized and sham-injected controls, suggesting that the cellular immune response was a factor in conferring resistance to fatal infection by AcMNPV-hsp70/lacZ.