FIG. 4.

Neither wild-type nor mutant bacteria trigger a respiratory burst in primary human monocytes and MDM. (A) Superoxide production by resting MDM (Un) or cells stimulated with 200 nM PMA, zymosan (Zymo., MOI of 5:1), LVS, the carB mutant, or formalin-killed LVS (FK-LVS) (each at an MOI of 50:1) was quantified at 30-s intervals over 60 min at 37°C as lucigenin CL. Data indicate the means of the results for triplicate samples from one experiment that is representative of four. (B) NBT staining of MDM demonstrates accumulation of superoxide inside zymosan phagosomes (Zym, black arrows) but not compartments containing LVS or the carB mutant (white arrowheads). Data shown are representative of three independent experiments. (C) Confocal sections of MDM stained to show gp91^phox/p22^phox heterodimers in green and LVS in red. Left panel, distribution of gp91^phox/p22^phox in uninfected MDM. Arrows indicate the plasma membrane, and the arrowhead indicates the biosynthetic-secretion pathway. Middle panel, MDM infected with zymosan for 15 min. Arrows indicate phagosomes. Right panel, MDM infected with LVS (red) for 15 min. Arrows indicate LVS phagosomes. (D) Superoxide production by resting monocytes (Un) or cells stimulated with zymosan (Zymo.; MOI, 5:1), LVS, the carA or carB mutant, or the single-iglC mutant (each at an MOI of 50:1) was quantified over 60 min at 37°C as lucigenin CL. Data indicate the means of the results for triplicate samples from one experiment that is representative of four. (E) Same as described for panel D except that monocyte ROS were detected by using luminol. Un, untreated.