Effect of Porcine Reproductive and Respiratory Syndrome Virus on Porcine Alveolar Macrophage Function as Determined Using Serial Analysis of Gene Expression (SAGE)

Abstract

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major pathogen of swine worldwide and causes considerable economic loss. The main target of infection is the porcine alveolar macrophage (PAM). Infection of PAMs by PRRSV causes significant changes in their function by mechanisms that are not understood. We have employed Serial Analysis of Gene Expression (SAGE) to examine the global expression of genes in PRRSV-infected PAMs. Total cellular RNAwas prepared from in vitro mock-infected and PRRSV strain VR-2332-infected PAMs at 0, 6, 12, 16 and 24 hours after infection, and subjected to SAGE analysis to obtain > 100,000 tags per time point. These sequences were processed to account for sequencing error before generating tag:count lists. These lists were deposited into a modified Identitag database for mapping to porcine and PRRSV genes. Identified unique mRNAtags were analyzed for their identity and relative abundance. Examination of the SAGE data indicated that there were changes in gene expression occurring in the PRRSV-infected PAMs over time post-infection. More than 400 unique tags with significantly altered expression levels were identified (p < 0.01 with Bonferroni correction). The validity and kinetics of expression of SAGE identified genes were evaluated using real-time RT-PCR.