Fig. 2: The cell envelope-associated LukAB is an active toxin.

a Intoxication of PMNs with cell-free bacterial lysate (Lysostaphin + ) or media control (Lysostaphin -). In the last group, anti-LukA antibodies (Anti-LukA + ) were added into the culture to neutralize LukAB activity. PMN viability was measured by LDH release after 2 h. Bars indicate mean ± SEM of 4 blood donors. ****p < 0.0001 by two-way ANOVA with Tukey’s multiple comparison test. b Infection of PMNs with WT or ∆lukAB USA300 ± 100 µg/ml chloramphenicol (Cm) at multiplicity of infection (MOI) = 100. PMN viability was measured by LDH release after 1 h. Bars indicate mean ± SEM of 12 blood donors. ***p = 0.0005, ****p < 0.0001 by RM one-way ANOVA with Tukey’s multiple comparison test. c Infection of PMNs with a leukocidin-null isogenic mutant strain complemented by hemin-inducible lukAB or pvl. The strains were pre-exposed ± 2 µM hemin before the infection. PMNs were infected without hemin, with 40 µg/ml tetracycline at MOI = 25. PMN viability was measured by LDH release after 1 h. Bars indicate mean ± SEM of 8 blood donors. **p = 0.0057 by RM one-way ANOVA with Tukey’s multiple comparison test. d Infection of PMNs with WT or ∆lukAB USA300 from the exponential phase (3 h), early stationary phase (5 h), and late stationary phase (24 h) + 100 µg/ml chloramphenicol (Cm) at MOI = 100. PMN viability was measured by LDH release after 1 h. Bars indicate mean ± SEM of 6 blood donors. p-values: WT-3h vs ∆lukAB-3h, 0.0003; WT-3h vs WT-5h, 0.0014; WT-5h vs ∆lukAB-5h, 0.0081; WT-5h vs WT-24h, 0.0076; WT-24h vs ∆lukAB-24h, 0.9999, determined by RM one-way ANOVA with Tukey’s multiple comparison test. Source data are provided as a Source Data file.