(A) T3SS1 cytotoxicity toward HT-29 and SLC35B2 mutant cells without pretreatment (PBS), preincubated then washed (coating) with GAGs (500µg/ml) (sulfated heparin (HS); dermatan sulfate (DS); chondroitin sulfate (CSA), or non-sulfated: hyaluronic acid (HA)), or infected in the presence of heparin (500µg/ml) (blocking). (B) Adherence of T3SS- V. parahaemolyticus to HT-29 and SLC35B2 mutant cells in presence of 500µg/ml sulfated or non-sulfated GAGs. (C) Effect of GAGs on resistance of HT-29 cells to T3SS1 and T3SS2 killing. (D) Survival kinetics of host cells of varying genotypes following infection with T3SS1+ and T3SS-deficient V. parahaemolyticus. (E) Survival kinetics of HT-29 and SLC35B2 mutant cells during infection with either T3SS1+ or T3SS1+
V. parahaemolyticus expressing the Afa-I adhesin or following infection initiated with centrifugation of V. parahaemolyticus onto host cells (spin). (F) Translocation of the T3SS1 effector VopQ fused to adenylate cyclase (CyA) into different host cells by V. parahaemolyticus after a 20-minute infection. Translocation into SLC35B2 cells was evaluated with and without centrifugation (spin) to enhance bacterial attachment. Data are mean with SEM (n=3). P values (* < 0.01, ** < 0.001, *** < 0.0001) are based on one-way ANOVA with Dunnet post test correction.