Figure 3. High lytic activity of ClyR.

(A) Time-kill curves of ClyR against S. dysgalactiae ATCC 35666. Bacterial cells were washed once with PBS, treated with 25 μg/ml ClyR, and the change of OD₆₀₀ (right Y-axis) were monitored by a microplate reader at 37 °C for 30 min. Triangles: ClyR; Circles: PBS controls. In parallel, the viable cell numbers (squares, left Y-axis) were calculated by plating onto BHI agar plates at different time points. (B) Dose-dependent lytic efficacy of ClyR against S. agalactiae S12, S. dysgalactiae ATCC 35666, or an equal mixture of both strains in market pasteurized milk at 30 °C for 1 h. (C) TEM images of S. dysgalactiae ATCC 35666 cells exposed to ClyR. Bar sizes: 500 nm. (D) Time-killing efficacy of ClyR (40 μg/ml) against S. agalactiae S12, S. dysgalactiae ATCC 35666, or an equal mixture of both strains in market pasteurized milk at 30 °C. (E) Lytic efficacy of ClyR (40 μg/ml) in pasteurized cow’s milk against S. agalactiae S12, or S. dysgalactiae ATCC 35666 at 30 °C for 1 h. Fresh cow milk samples A, B, and C were taken from healthy cows, and samples D and E were from mastitic cows. (F) Comparison of the activity of 0.89 μM ClyR or PlyCAC against various strains. (G) Comparison of the activity of 0.89 μM ClyR or PlyGBS-180 against S. dysgalactiae ATCC 35666. (H) Efficacy of PlyGBS-180 against S. dysgalactiae ATCC 35666 in market pasteurized milk at 30 °C for 60 min.