the hemolymph and tracheal system to invade and infect other organs and tissues of the insect larva. Replication of the Gammabaculoviruses is restricted to the midgut (144). At the end of the infection virions are occluded into OBs, which are released into the environment from the insect body upon death. In the OB form baculoviruses can persist in the environment for many years. For some baculoviruses, notably AcMNPV, the replication cycle can also be completed in cell culture using BVs as inoculum and this has greatly enhanced our current understanding of the cell biology and genetics of baculovirus infections (for more details (279)).
Baculoviruses contain a double-stranded, circular and superhelical DNA molecule, which replicates in the cell nucleus. After synthesis the DNA is packaged in a number of proteins to form nucleocapsids. One or more nucleocapsids are wrapped in an envelope, which is de novo formed in the nucleus of infected cells. Single or multiple virion packages are occluded in GVs and NPVs, respectively. The size of baculovirus genomes depends on the species and ranges from 80 to 180 kilobasepairs (kbp), hence encompassing a variable numbers of ORFs (266). Gene homology, gene content and gene location can be used to construct reliable phylogenetic trees to show the relatedness among baculoviruses (99).
The transcription of baculoviruses occurs in a cascaded fashion and four classes of transcripts are discriminated. The immediate early (IE) transcripts are made by host RNA polymerases, a process independent of de novo protein synthesis. Delayed early (DE) transcripts require translation of IE viral transcripts for their synthesis. Late (L) transcripts are expressed after the onset of DNA replication and very late (VL) transcripts are those that are still expressed very late after infection, sometimes at very high levels (polyhedrin, p10). Baculovirus late and very late transcription occurs via a virus-encoded RNA polymerase, which is α-amanitin insensitive. Baculovirus genes are hence categorized as immediate-early, delayed-early, late and very late genes. Baculovirus transcripts may have 5'and 3' co-terminal ends and hence may overlap in sequence and time of expression. A canonical baculovirus transcription initiation motif (TAAG) is present in the promoter region of L and VL genes, whereas a more common motif (CAGT) is often associated with early baculovirus gene expression (recent review (213)).
The fact that for replication in cell culture only BVs are required and the ODV phenotype is dispensable allowed the development of the baculovirus expression system for production of recombinant protein in insect cells (254). Baculovirus VL genes are highly expressed but not required for virus propagation in cell culture, enabling their replacement with foreign genes. The promoters of the polyhedrin and p10 genes are extensively used as cassettes to drive the expression of single or multiple foreign genes in a baculovirus background. The development of an AcMNPV bacmid greatly facilitated the functional analysis of baculovirus genes, since it not only simplified the engineering of baculovirus expression vectors (172), but also made the construction of (knock-out) mutants much easier. The deletion of the cathepsin and the chitinase genes for instance, has improved the integrity of secreted recombinant proteins (120). The insertion of genes for the modification of glycoproteins in the Golgi system, has allowed the production of complex, mammalian-like glycoproteins in insect cells (115). AcMNPV can also be used as a delivery vector for mammalian cells and as gene therapy vector, either as a gene carrier or as a production system for other gene therapy vectors such as adeno-associated viruses (108, 174, 244). The baculovirus insect cell expression system is still being tailored and optimised to meet the demands of both the scientist as well as the commerce.
AcMNPV was isolated in 1969 by the late Dr. Patrick V. Vail and colleagues from a single virosed insect larva near Riverside (264). The insect was assigned a