One hundred Hampshire by Duroc crossbred pigs (HD) and 100 NE Index line pigs (I) were infected with porcine reproductive and respiratory syndrome virus (PRRSV) and evaluated for resistance/susceptibility. Controls (100/line) were uninfected littermates to infected pigs. Viremia (V), weight change (WT?), and rectal temperature at 0, 4, 7, and 14 days post-infection (dpi) were recorded. Lung, bronchial lymph node (BLN), and blood tissue were collected at necropsy (14 dpi). Infected pigs were classified as low or high responders to PRRSV based on the first principal component (PC) from principal component analyses of all variables. Low responders to PRRSV (low PRRSV burden) and their uninfected littermates were assigned to low (L) class. High responders to PRRSV (high PRRSV burden) and their uninfected littermates were assigned to high (H) class. Infected pigs in the L-class had high WT?, low V, and few lung lesions; H-class pigs had low WT?, high V, and many lung lesions. RNA was extracted from lung and BLN tissue of the seven highest and seven lowest responders per line and from each of their control littermates. A control reference design was used and cDNA from each reference sample tissue was prepared from pooled RNA extracted from two control pigs from each line whose infected littermates had a PC value of 0. Design variables in data analyses were line (I vs HD), class (H vs L), treatment (infected vs uninfected controls), and slide/pig as error. Overall design: A gene expression study was conducted with 56 pigs from a PRRSV-infection experiment involving a total of 400 pigs. Two hundred pigs from the NE Index line (I), selected 20 generations for increased litter size, and 200 pigs from a commercial Hampshire x Duroc (HD) cross selected for lean growth, were used. Two pigs of the same sex from each available litter, representing 83 sires and 163 dams, were selected at random for the experiment. A total of 200 pigs were infected with PRRSV and 200 uninfected littermates served as controls. The experiment was conducted in two replicates within each of two seasons with 50 pigs per breed in each year/season/replicate. Pigs were housed in two isolation rooms at the University of Nebraska Animal Research Facility of the Veterinary and Biomedical Sciences Department. Each room contained two pens of 12-13 pigs per pen, with Line I pigs in one pen and Line HD pigs in the other pen. One room was randomly assigned for treatment and pigs in it were inoculated intranasally with 105 CCID50 of PRRSV strain 97-7985. Application rate was 1cc per nostril. Littermates, serving as controls to the infected pigs, were placed in the second room. Phenotypic data included viremia (V) from serum samples collected at 4, 7, and 14 days post-infection (dpi), changes in weight (WT?) and rectal temperature from 0 to 4, 4 to 7, and 7 to 14 dpi, lung and bronchial lymph node viremia from tissue collected at necropsy at day 14, and severity of lung lesions. Blood serum at day 0 before infection was collected and stored. Samples were collected at 4, 7, and 14 dpi to monitor changes during and shortly after the acute phase of viral infection. Uninfected HD pigs gained more and had higher rectal temperature from 0 to 14 dpi than uninfected I pigs, whereas infected I pigs gained more and had lower temperature than infected HD pigs. Viremia (CCID50/mL, cell culture infectious dose 50%/mL) was also greater in HD than I pigs at 4, 7, and 14 dpi. Genetic line differences in lung and lymph viremia levels were not significant, but tended to be greater in HD than I pigs. Based on these results, the current microarray experiment was designed to determine whether expression of genes differed between pigs in the tails of the PRRSV response distribution. Phenotypic data for infected pigs were subjected to principal component (PC) multivariate procedures to identify 28 pigs, seven pigs within each line in the outermost tails of the distributions of viral response variables. The first PC eigenvector, which accounted for 27% of the variation, was used to rank pigs. The 28 control littermates to each of these pigs were also selected, resulting in 56 pigs used in the gene expression experiment. Infected pigs in the right tail of the PC distribution were considered to be susceptible to PRRSV. Those pigs and their control littermates formed the H class (high PRRSV burden). Pigs in the left tail were considered to be tolerant or resistant. They and their control littermates formed the L class (low PRRSV burden). A 2x2x2 factorial treatment design was utilized for the gene expression experiment. Design effects included class (H and L), line (I and HD), and treatment (infected and uninfected), with seven pigs in each of the eight categories.