SUMMARYThe fl-galactosidase gene (lacZ) of Escherichia coli was inserted in phase with the coding sequence of the Autographa californica nuclear polyhedrosis virus (AcMNPV) late-expressed Mr 10000 (pl0) gene. The fusion gene was inserted into the AcMNPV genome by cotransfection of a recombinant plasmid pAcR159Z, consisting of the EcoRI P fragment-containing pBR325-derived plasmid pAcR159 and the lacZ insert inthe p 10 gene, and wild-type AcMNPV DNA. Infection of Spodopterafrugiperda cells by the resulting recombinant AcMNPV/p 10Z-2 showed high level expression of a p 10-lacZ fusion protein, but no synthesis of pl0. Therefore, the pl0 gene is dispensable for virus replication and the pl0 promoter is effective in driving the expression of foreign genes. Cells infected with AcMNPV/pl0Z recombinants resembled those infected with wild-type AcMNPV in the amounts of polyhedrin synthesized and polyhedra formed, although pl0 was absent. The nucleus and cytoplasm of AcMNPV/pl0Z-2-infected cells lacked the fibrous structures that are associated with pl0 in wild-type AcMNPV-infected cells. Instead, large granular structures were observed that were found by immunogold labelling to contain the lacZ gene product. The electron-dense 'spacers', thought to be precursors of the polyhedron membrane, were absent from cells infected by the recombinant virus and the polyhedra did not have a membrane. The recombinant AcMNPV/pl0Z-2 was at least twice as virulent for second instar S. exigua larvae than was wild-type AcMNPV. The increased virulence of the recombinant is an important property for the control of insects.

SUMMARYWe have investigated the behaviour of coat protein deletion mutants of the geminivirus African cassava mosaic virus (ACMV) when introduced into Nicotiana benthamiana by agroinoculation. In dividing callus tissue, replicating mutant DNA A, in the absence of DNA B, remained subgenomic in size. However, systemic infection of plants was associated with the rapid reversion of the deletion mutants to native component size, as happened when the mutants were introduced into the host by more conventional mechanical inoculation procedures. The results contrast with those reported for tomato golden mosaic virus (TGMV) for which similar mutants, agroinoculated into N. benthamiana, remained subgenomic. The results indicate that the inoculation route is not responsible for the different behaviour of ACMV and TGMV mutants and illustrate a significant difference between the interaction of these otherwise closely related geminiviruses with a common host that might be exploited for the investigation of the determinants of DNA replication and spread.

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