Page Banner

United States Department of Agriculture

Agricultural Research Service

Research Project: GENE EXPRESSION IN LYMPH NODES OF PRRSV-INFECTED PIGS Title: Immune transcriptome response to PRRSV infection

Authors
item MILLER, LAURA
item HARHAY, GREGORY
item LAGER, KELLY
item Kehrli Jr, Marcus

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: May 11, 2011
Publication Date: June 4, 2011
Citation: Miller, L.C., Harhay, G.P., Lager, K.M., Kehrli, Jr., M.E. 2011. Immune transcriptome response to PRRSV infection [abstract]. XIIth International Nidovirus Symposium. p. 79.

Technical Abstract: Porcine reproductive and respiratory syndrome virus (PRRSV) is a major pathogen of swine. Here we identified and compared gene expression changes in tracheobronchial lymph nodes (TBLN) following viral infection using Digital Gene Expression Tag Profiling (DGETP). Pigs were infected with 1 x 105 cell culture infectious dose 50% per pig of PRRSV (strain SDSU73), porcine circovirus type 2 (PCV2; group 2), or swine influenza virus (SIV; A/SW/OH/511445/2007). Pigs were allotted to one of 4 treatment groups: sham inoculated control, PRRSV-challenge, PCV2-challenge, or SIV-challenge. On 0 dpi pigs received an intranasal challenge with 2 ml of either sham or virus inoculum. Control pigs were sham inoculated with tissue culture supernatant. Five pigs from each group were euthanized and necropsied on 1, 3, 6, and 14 dpi. Body temperatures of pigs euthanized on 14 dpi were recorded daily. At necropsy, lungs were scored for gross lesions. TBLN were homogenized and aliquots used for RNA extraction or immunophenotyped by flow cytometry analysis and cytokine expression.. Total RNA was pooled for each group and timepoint to make 16 libraries, for analysis by DGETP using the whole-genome expression analysis platform (Illumina Technologies). Data underwent image analysis, base calling, and standard filtering to generate a list of sequence tags and counts. Multidimensional statistical tests and clustering analysis identified significant changes in tag abundance. PRRSV infection reduced the unique tag sequences (i.e., transcriptome diversity) in the TBLN transcriptome at 1, 3, 6 and 14 dpi to 55.7%, 69.3%, 56.4% and 55.5% of control TBLN. PCV2 infection increased the unique tag sequences to 95.0%, 111.6%, 148.7% and 106.8% of controls where as, SIV induced a more modest reduction in unique tag sequences at 87.1%, 67.6%, 100.7% and 70.9% of the respective control transcriptome at 1, 3, 6 and 14 dpi. Tags were annotated with available transcript information from7804 swine RefSeq sequences and 240420 HarvardGI Accessions. GenBank and Refseq virus sequences were used to determine and subtract viral tag counts. For select genes of interest, transcript changes were validated by real-time RT-PCR, and transcripts with changes in abundance were mapped to known metabolic, signaling and other pathways/networks. Gaining insight into how PRRSV causes disease may aid development of more efficacious vaccines.

Last Modified: 9/10/2014
Footer Content Back to Top of Page