title: The real-time-based assessment of the microbial killing by the antimicrobial compounds of neutrophils.
abstract: A recombinant Escherichia coli K-12 strain, transformed with a modified bacterial luciferase gene (luxABCDE) from Photorhabdus luminescens, was constructed in order to monitor the activity of various antimicrobial agents on a real-time basis. This E. coli-lux emitted, without any addition of substrate, constitutive bioluminescence (BL), which correlated to the number of viable bacterial cells. The decrease in BL signal correlated to the number of killed bacterial cells. Antimicrobial activity of hydrogen peroxide (H(2)O(2)) and myeloperoxidase (MPO) was assessed. In high concentrations, H(2)O(2) alone had a bacteriocidic function and MPO enhanced this killing by forming hypochlorous acid (HOCl). Taurine, the known HOCl scavenger, blocked the killing by MPO. When E. coli-lux was incubated with neutrophils, similar killing kinetics was recorded as in H(2)O(2)/MPO experiments. The opsonization of bacteria enhanced the killing, and the maximum rate of the MPO release from lysosomes coincided with the onset of the killing.
authors.affiliation: Laboratory of Immunochemistry, Department of Biochemistry and Food Chemistry, University of Turku, Vatselankatu 2, 20014 Turku, Finland. janato@utu.fi
authors.forename: J T
authors.initials: JT
authors.lastname: Atosuo
authors.forename: E-M
authors.initials: EM
authors.lastname: Lilius
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D058491:Bacterial Load; D015169:Colony Count, Microbial; D048168:Escherichia coli K12 / Q000737:chemistry / Q000187:drug effects* / Q000276:immunology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006997:Hypochlorous Acid / Q000737:chemistry; D007700:Kinetics; D049453:Luciferases, Bacterial / Q000737:chemistry / Q000235:genetics; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics; D008247:Lysosomes / Q000737:chemistry / Q000276:immunology / Q000382:microbiology; D050296:Microbial Viability; D009504:Neutrophils / Q000737:chemistry* / Q000276:immunology; D009195:Peroxidase / Q000494:pharmacology; D010587:Phagocytosis; D020637:Photorhabdus / Q000201:enzymology / Q000235:genetics; D013654:Taurine / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D008164:Luminescent Proteins; D013654:Taurine; D006997:Hypochlorous Acid; D006861:Hydrogen Peroxide; D009195:Peroxidase; D049453:Luciferases, Bacterial