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Virus replication can cause extensive rearrangement of host cell cytoskeletal and membrane compartments leading to the “cytopathic effect” that has been the hallmark of virus infection in tissue culture for many years. Recent studies are beginning to redefine these signs of viral infection in terms of specific effects of viruses on cellular processes. In this chapter, these concepts have been illustrated by describing the replication sites produced by many different viruses. In many cases, the cellular rearrangements caused during virus infection lead to the construction of sophisticated platforms in the cell that concentrate replicase proteins, virus genomes, and host proteins required for replication, and thereby increase the efficiency of replication. Interestingly, these same structures, called virus factories, virus inclusions, or virosomes, can recruit host components that are associated with cellular defences against infection and cell stress. It is possible that cellular defence pathways can be subverted by viruses to generate sites of replication. The recruitment of cellular membranes and cytoskeleton to generate virus replication sites can also benefit viruses in other ways. Disruption of cellular membranes can, for example, slow the transport of immunomodulatory proteins to the surface of infected cells and protect against innate and acquired immune responses, and rearrangements to cytoskeleton can facilitate virus release.

Stress signaling from mitochondria and the endoplasmic reticulum (ER) leads to the induction of the proapoptotic transcription factor CHOP/GADD153. Many viruses use the ER as a site of replication and/or envelopment, and this activity can lead to the activation of ER stress and apoptosis. African swine fever virus (ASFV) is assembled on the cytoplasmic face of the ER and ultimately enveloped by ER membrane cisternae. The virus also recruits mitochondria to sites of viral replication and induces the mitochondrial stress protein hsp60. Here we studied the effects of ASFV on the induction of CHOP/GADD153 in infected cells. Interestingly, unlike other ER-tropic viruses, ASFV did not activate CHOP and was able to inhibit the induction of CHOP/GADD153 by a number of exogenous stimuli.

African swine fever virus (ASFV) is a large double-stranded DNA virus that replicates in discrete areas in the cytosol of infected cells called viral factories. Recent studies have shown that assembling virions acquire their internal envelopes through enwrapment by membranes derived from the endoplasmic reticulum (ER). However, the mechanisms that underlie the formation of viral factories and progenitor viral membranes are as yet unclear. Analysis of the published genome of the virus revealed a conserved multigene family that encodes proteins with hydrophobic signal sequences, indicating possible translocation into the ER lumen. Strikingly, two of these genes, XP124L and Y118L , encoded proteins with KDEL-like ER retention motifs. Analysis of XP124L and Y118L gene product by biochemical and immunofluorescence techniques showed that the proteins were localized to pre-Golgi compartments and that the KEDL motif at the C terminus of pXP124L was functional. XP124L expression, in the absence of other ASFV genes, had a dramatic effect on the contents of the ER that was dependent precisely on the C-terminal sequence KEDL. The normal subcellular distribution of a number of proteins resident to this important, cellular organelle was drastically altered in cells expressing wild-type XP124L gene product. PXP124L formed unusual perinuclear structures that contained resident ER proteins, as well as proteins of the ER-Golgi intermediate compartment. The data presented here hint at a role for MGF110 gene product in preparing the ER for its role in viral morphogenesis; this and other potential functions are discussed.

African swine fever virus (ASFV) is a complex DNA virus and the aetiological agent of a domestic pig disease that can assume different clinical forms, ranging from an acute haemorrhagic disease to unapparent infections. The complexity of the immune response has impaired the development of an effective vaccine against ASFV infection. Previous work has established that immunization of pigs with attenuated replication competent ASFV strains can induce good levels of protection against lethal challenge with virulent strains. However in some pigs adverse clinical signs occur following immunization. On the other hand, due to the highly acute nature of the disease following infection with the virulent strains, it is clear that the porcine innate response is insufficient to prolong the life of the animal until the serological and cellular adaptive immune responses provide immediate protection and, importantly, the generation of the immunological memory crucial to vaccine development. The type I interferon (IFN) response is the first line of innate immunity against viral infection. Most, if not all viruses have to overcome this host defense before they can establish productive infection. We are currently characterizing the induction of type I IFN in primary porcine macrophages infected with strains of different virulence. A better understating of how the IFN response is modulated by these different viruses, will provide a rational basis for the development of novel vaccine strategies, for example, further attenuation of the virus by deleting genes involved in the evasion of the IFN host system is in progress.

African swine fever (ASF) is an acute haemorrhagic disease of domestic pigs for which there is currently no vaccine. We showed that experimental immunisation of pigs with the non-virulent OURT88/3 genotype I isolate from Portugal followed by the closely related virulent OURT88/1 genotype I isolate could confer protection against challenge with virulent isolates from Africa including the genotype I Benin 97/1 isolate and genotype X Uganda 1965 isolate. This immunisation strategy protected most pigs challenged with either Benin or Uganda from both disease and viraemia. Cross-protection was correlated with the ability of different ASFV isolates to stimulate immune lymphocytes from the OURT88/3 and OURT88/1 immunised pigs.

African swine fever virus (ASFV) encodes multiple copies of MGF360 and MGF530/505 gene families. These genes have been implicated in the modulation of the type I interferon (IFN) response. We investigated the effect of modulating the IFN response on virus attenuation and induction of protective immunity by deleting genes MGF360 (MGF360-10L, 11L, 12L, 13L, 14L) and MGF530/505 (MGF530/505-1R, 2R and 3R) and interrupting genes (MGF360-9L and MGF530/505-4R) in the genome of the virulent ASFV isolate Benin 97/1. Replication of this deletion mutant, BeninΔMGF, in porcine macrophages in vitro was similar to that of the parental virulent virus Benin 97/1 and the natural attenuated isolate OURT88/3, which has a similar deletion of MGF360 and 530/505 genes. Levels of IFN-β mRNA in macrophages infected with virulent Benin 97/1 isolate were barely detectable but high levels were detected in macrophages infected with OURT88/3 and intermediate levels in macrophages infected with BeninΔMGF. The data confirms that these MGF360 and MGF530/505 genes have roles in suppressing induction of type I IFN. Immunisation and boost of pigs with BeninΔMGF showed that the virus was attenuated and all pigs (5/5) were protected against challenge with a lethal dose of virulent Benin 97/1. A short transient fever was observed at day 5 or 6 post-immunisation but no other clinical signs. Following immunisation and boost with the OURT88/3 isolate 3 of 4 pigs were protected against challenge. Differences were observed in the cellular and antibody responses in pigs immunised with BeninΔMGF compared to OURT88/3. Deletion of IFN modulators is a promising route for construction of rationally attenuated ASFV candidate vaccine strains.