Submitted to: Advances in Virology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 12/12/2010
Publication Date: 1/16/2011
Citation: Miller, L.C., Neill, J.D., Harhay, G.P., Lager, K.M., Laegreid, W.W., Kehrli, Jr., M.E. 2010. In-depth global analysis of transcript abundance levels in porcine alveolar macrophages following infection with porcine reproductive and respiratory syndrome virus. Advances in Virology. 2010:Article 864181. Available: http://downloads.hindawi.com/journals/av/2010/864181.pdf. Interpretive Summary: The pig respiratory virus, porcine reproductive and respiratory syndrome virus (PRRSV), causes highly significant losses to the swine industry worldwide. The ability of the virus to persist in its host shows that it has mechanisms to evade host immune responses. This study examined the effect of porcine reproductive and respiratory virus (PRRSV) on how genes are expressed in porcine alveolar macrophages (PAMs). PAMs were chosen for this study because they are the primary targets of infection by PRRSV. Serial analysis of gene expression (SAGE) was used because it allowed us to look at all genes expressed in these cells. We determined the normal levels of gene expression in normal, non-infected PAMs and then compared this to gene expression levels in PRRSV-infected PAMs at several time points after infection. It is well established that many pathogens interfere with expression of specific genes that act to protect the host and clear the infection. Of particular interest was the minimal expression of genes that are involved in attracting other immune cells to the area of the infection. Additionally, there was no response by genes that cause inflammation. This is the first comprehensive study to show the inhibition of an immune response in PAMs by PRRSV. Results have also given us clues to the mechanism(s) behind this inhibition. There are specific cellular proteins that control expression of protective genes and future studies will look at how this virus may be inhibiting their function. This may possibly lead to a means for developing a vaccine that may act to limit or end an active PRRSV infection by restoring the cell’s natural protective mechanisms.
Technical Abstract: Porcine reproductive and respiratory syndrome virus (PRRSV) is a major pathogen of swine worldwide and causes considerable economic loss. Infection of the primary target cells, porcine alveolar macrophages (PAMs), by PRRSV causes significant changes in their function by mechanisms that are not understood. Identifying specific pathways that associate with variation in PRRSV replication and macrophage function may lead to novel gene targets for the control of PRRSV infection. Serial Analysis of Gene Expression (SAGE) libraries were constructed from in vitro mock-infected and PRRSV strain VR-2332-infected PAMs at 0, 6, 12, 16 and 24 hours post-infection. Each SAGE library was sequenced to obtain >95,000 tags per time point. Tags were mapped to transcripts and genes by exact regular expression matching to sequences in GenBank, Harvard Gene Index, and the Pig Expression Database (Japan). Examination of the SAGE data indicated that there were changes in transcript abundance occurring in the PRRSV-infected PAM over time post-infection. More than 590 unique tags with significantly altered transcript abundance levels were identified (p<0.01 with Bonferroni correction). The validity and kinetics of transcript abundance of SAGE identified genes were confirmed using real-time RT-PCR. The most striking finding was that the transcript abundance of most of the identified genes involved in the innate immune response (including IL-8, CCL4, and IL-1Beta, genes whose transcript abundances are typically altered in response to other pathogens or insults) showed no or very little change at any time point following infection.