FIGURE 1.

NADase contributes to intracellular multiplication of GAS in endothelial cells. (A) NADase activity of A20, SF370, and NZ131. NADase activity was determined by co-incubation with NAD⁺ and expressed as relative percentage compared to medium alone. (B) HMEC-1 cells were infected with serotype M1 SF370 and A20 (M.O.I. = 10), or M49 NZ131 (M.O.I. = 1) for 30 min, and treated with gentamicin to kill extracellular bacteria. Intracellular viable bacteria were counted by CFU-based assays. (C) NADase activity was determined by co-incubation with NAD⁺ and expressed as relative percentage compared to medium alone. (D) Wild-type NZ131 and its isogenic strains infected HMEC-1 cells at M.O.I. of 1 and intracellular viable bacteria were counted by CFU-based assays. (E) NADase activities of NZ131, nga mutant, ifs-overexpressing, and vector control strains were determined by co-incubation with NAD⁺ and expressed as relative percentage compared to medium alone. (F) HMEC-1 cells were infected with ifs-overexpressed NZ131 at M.O.I. of 1 and intracellular viable bacteria were counted by CFU-based assays. The data represent the means ± SEM of at least three independent experiments. ^∗∗∗p < 0.001 (one- or two-way ANOVA).