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. 2023 Aug 3;13:12618. doi: 10.1038/s41598-023-39613-5

Figure 2.

Figure 2

Colistin resistant Kp209_CSTR has been sensitized to MAC-mediated killing. (a) Inner membrane permeabilization of Kp209, Kp257 and Kp040, and their colistin resistant derived strains Kp209_CSTR, Kp257_CSTR and Kp040_CSTR in the presences of 10% NHS, 10% NHS in which C5 conversion was inhibited by addition of 20 µg/ml OMCI and 20 µg/ml Eculizumab (C5 inhibition), or 10% heat inactivated NHS (HiNHS). Bacteria were incubated at 37 °C in the presence of 1 µM SYTOX green nucleic acid stain, and inner membrane permeabilization (SYTOX fluorescence intensity) was detected every 2 min for 90 min in a microplate reader. (b) Survival on plate of Kp209, Kp257 and Kp040, and their colistin resistant derived strains Kp209_CSTR, Kp257_CSTR, and Kp040_CSTR after 90 min incubation in 10% NHS at 37 °C. Survival data is normalized to CFU counts in conditions where the terminal complement pathway was blocked by inhibiting C5 conversion (20 µg/ml OMCI and 20 µg/ml Eculizumab). (c) Inner membrane permeabilization of Kp257, Kp257_CSTR, Kp040 and Kp040_CSTR in the presence of 50% NHS, or 50% HiNHS. Bacteria were incubated at 37 °C in the presence of 1 µM SYTOX green nucleic acid stain, and inner membrane permeabilization (SYTOX fluorescence intensity) was detected every 6 min for 90 min in a microplate reader. (d) Survival on plate of Kp257, Kp257_CSTR, Kp040 and Kp040_CSTR after 90 min incubation in 50% NHS at 37 °C. Survival data was normalized to CFU counts in 50% HiNHS. (a–d) Data represent mean ± standard deviation of three independent experiments.