Spread of recombinant Autographa californica nucleopolyhedrovirus in various tissues of silkworm Bombyx mori determined by real-time PCR

Abstract

A cassette harboring luciferase reporter driven by Bombyx mori A3 promoter was transferred to the bacmid AcΔEGT to generate the recombinant virus AcNPVA3Luc (where Ac represents Autographa californica, NPV represents nucleopolyhedrovirus, and A3Luc represents the firefly luciferase reporter cassette driven by the A3 promoter). Recombinant baculovirus was injected into the hemocoele of newly ecdysed fifth instar larvae of the silkworm. The infection of virus in various silkworm tissues was determined by real-time PCR. The profile of viral infection showed that the copy number of recombinant AcNPV (rAcNPV) increased the fastest in the hemocyte, followed by the fat body, Malpighian tubule, middle gut, and silk gland. Detecting in nonpermissive strain silkworm showed that there was no significant difference in the entry of rAcNPV into all tested tissues. The difference in viral infection reflected mainly the big difference in replication of rAcNPV in various tissues of silkworm larvae. Real-time quantitative RT–PCR showed that it was due to the different expression of genes involved in viral DNA replication.