Porcine reproductive and respiratory syndrome virus (PRRSV) is an arterivirus that emerged in the late 1980s in both Europe and North America as the causative agent of porcine reproductive and respiratory syndrome (PRRS), now the most important disease of swine worldwide. Despite extensive characterization of PRRSV proteins by direct analysis and comparison with other arteriviruses, determinants of virulence, pathogenesis and protective immune recognition remain poorly understood. Thus, we hypothesized that additional ORFs are present in the PRRSV genome that may contribute to its biological properties, and so we screened highly purified virions of strain VR2332, the prototype type 2 PRRSV, for evidence of novel polypeptides. A 51 aa polypeptide was discovered that is encoded by an alternative ORF of the subgenomic mRNA encoding the major envelope glycoprotein, GP5, and which is incorporated into virions. The protein, referred to as ORF5a protein, is expressed in infected cells, and pigs infected with PRRSV express anti-ORF5a protein antibodies. A similar ORF is present as an alternative reading frame in all PRRSV subgenomic RNA5 genes and in all other arteriviruses, suggesting that this ORF5a protein plays a significant role in arterivirology. Its discovery also provides a new potential target for immunological and pharmacological intervention in PRRS.

The immunology of porcine reproductive and respiratory syndrome virus (PRRS) begins with an initial encounter of PRRSV with the pig. Regardless of the route of entry of PRRSV--via inhalation, intramuscular vaccination, insemination, or other routes--productive infection occurs predominately in alveolar macrophages of the lung. Thus, innate responses of the lung and the alveolar macrophage comprise the initial defense against PRRSV. The virus appears not to elicit innate interferon and cytokine responses characteristic of other strongly immunogenic viral pathogens, and its effects are consistent with induction of a weak adaptive immune response. Humoral and cell-mediated immunity is induced in due course, and results in clearance of virus from the circulation but not from lymphoid tissues, where the infection becomes persistent. Subsequent reexposure to PRRSV elicits an anamnestic response that is partially to completely protective. Within this unconventional picture of anti-PRRSV immunity lie a variety of unresolved issues, including the nature of protective immunity within individual pigs and among pigs in commercial populations, the efficacy of protective immunity against genetically different PRRSV isolates, the effects of developmental age, sex, genetics, and other host factors on the immune response to PRRSV, and the possible suppression of host immunity to other pathogens.

Background: The domestic pig is known as an excellent model for human immunology and the two species share many pathogens. Susceptibility to infectious disease is one of the major constraints on swine performance, yet the structure and function of genes comprising the pig immunome are not well-characterized. The completion of the pig genome provides the opportunity to annotate the pig immunome, and compare and contrast pig and human immune systems. Results: The Immune Response Annotation Group (IRAG) used computational curation and manual annotation of the swine genome assembly 10.2 (Sscrofa10.2) to refine the currently available automated annotation of 1,369 immunity-related genes through sequence-based comparison to genes in other species. Within these genes, we annotated 3,472 transcripts. Annotation provided evidence for gene expansions in several immune response families, and identified artiodactyl-specific expansions in the cathelicidin and type 1 Interferon families. We found gene duplications for 18 genes, including 13 immune response genes and five non-immune response genes discovered in the annotation process. Manual annotation provided evidence for many new alternative splice variants and 8 gene duplications. Over 1,100 transcripts without porcine sequence evidence were detected using cross-species annotation. We used a functional approach to discover and accurately annotate porcine immune response genes. A co-expression clustering analysis of transcriptomic data from selected experimental infections or immune stimulations of blood, macrophages or lymph nodes identified a large cluster of genes that exhibited a correlated positive response upon infection across multiple pathogens or immune stimuli. Interestingly, this gene cluster (cluster 4) is enriched for known general human immune response genes, yet contains many un-annotated porcine genes. A phylogenetic analysis of the encoded proteins of cluster 4 genes showed that 15% exhibited an accelerated evolution as compared to 4.1% across the entire genome. Conclusions: This extensive annotation dramatically extends the genome-based knowledge of the molecular genetics and structure of a major portion of the porcine immunome. Our complementary functional approach using co-expression during immune response has provided new putative immune response annotation for over 500 porcine genes. Our phylogenetic analysis of this core immunome cluster confirms rapid evolutionary change in this set of genes, and that, as in other species, such genes are important components of the pig’s adaptation to pathogen challenge over evolutionary time. These comprehensive and integrated analyses increase the value of the porcine genome sequence and provide important tools for global analyses and data-mining of the porcine immune response.

Porcine reproductive and respiratory syndrome virus (PRRSV) is the most important infectious disease agent of pigs worldwide, causing reproductive failure in pregnant sows and respiratory problems in nursing and growing pigs. PRRSV infection is characterized by a prolonged viremia of 30 or more days and an extended persistent infection of lymphoid tissues. To better understand the immunological basis for prolonged acute and persistent PRRSV infection, we have examined the cell-mediated immune (CMI) response throughout the course of infection and compared the results to the local distribution and abundance of PRRSV in infected tissues. PRRSV-specific T cells, enumerated by gamma interferon enzyme-linked immunospot assay, did not appear until 2 weeks after PRRSV inoculation, and their abundance exhibited substantial variation over time and among animals. In all cases the T-cell response was transient. High levels of viral RNA were present in lymphoid tissues of all animals in the acute phase of infection. Viral loads were decreased 1,000-fold or more in persistent infections, with the primary sites of persistence being tonsil, sternal lymph node, and inguinal lymph node. The abundance of virus-specific T cells in either acutely or persistently infected animals was highly variable and showed no correlation to the level of virus in lymphoid tissues. No significant difference in antigen-specific T-cell abundance was observed in secondary lymphoid tissues in either acute or persistent infection except for tonsil, in which the number of responding cells was extremely low. CD4⁺- and CD8⁺-T-cell frequencies did not change after PRRSV infection, though a decrease in γδ T cells was observed. Macrophages, the permissive cell type for PRRSV, were present in various levels in all tissue preparations and were not in proportion to local virus load. These findings indicate that a weak CMI response contributes to prolonged PRRSV infection and suggests that PRRSV suppresses T-cell recognition of infected macrophages. Thus, the slow but eventual resolution of PRRSV infection may be dependent on limiting permissive macrophages and on innate immune factors.