FIG. 2.

Significant reductions occurred in NK-cell cytotoxicity despite a lack of significant changes in NK-cell populations following PRRSV and PRCV co-infection in pigs. (A) Percentages of CD3⁻CD4⁻CD8α⁺ lymphocytes (NK cells) in PBMCs of pigs were evaluated in mock-infected, PRRSV-infected, PRCV-infected, or dual-infected (PRCV/PRRSV) pigs on the indicated post-inoculation days (PID) by flow cytometric analysis. Each bar represents the mean ± SEM of NK cells from 4–8 pigs, and total numbers of pigs at each PRCV/PRRSV PID: −2/8 (n = 17); 2/12 (n = 18); 4/14 (n = 18); 8/18 (n = 20); 10/20 (n = 20); 14/24 (n = 21); and 21/31 (n = 21). (B) Percentages of NK-specific cytotoxicity were measured using pig PBMCs (effectors) harvested from mock-infected or infected pigs as described above against target cells (K-562). Effectors and targets at the different indicated ratios were cultured together, and the supernatants harvested were analyzed for amounts of LDH released using its substrate at 490 nm. Each line is from one pig, and each data point on the line is the mean of triplicate readings ± SEM at the respective E:T ratios tested. Altogether there were 19 pigs from one independent experimental trial (a total of four PIDs). A similar trend in NK-specific lysis was detected in another two independent experimental trials comprised of the same numbers of pigs. Statistical analyses were performed from the pooled results of all three independent experimental trials, and “a” denotes a statistically significant difference (p < 0.05) between the mock- and dual-infected groups; “b” denotes a statistically significant difference between the mock- and PRRSV-infected groups; “c” denotes a statistically significant difference between the mock- and PRCV-infected groups; and “d” denotes a statistically significant difference between the PRCV- and dual-infected groups, as analyzed by the nonparametric Kruskal-Wallis test.