title: Characterization of Staphylococcus aureus beta-toxin induced leukotoxicity.
abstract: One virulence determinant of Staphylococcus aureus is Beta-toxin, a 37 Kd magnesium-dependent sphingomyelinase C. This toxin lyses erythrocytes (RBCs) containing sphingomyelin in the outer lipid layer of their plasma membrane. Although membranes of both human polymorphonuclear leukocytes (PMNs) and lymphocytes (MNLs) contain small amounts of sphingomyelin, the effect of Beta-toxin on these cells remains controversial. The purpose of this study was to investigate the hemolytic activity of this toxin on RBCs of various species and determine the leukotoxic nature on several types of human leukocytes. One nanogram of Beta-toxin lysed 115,000 sheep erythrocytes (sRBCs) and 82,000 human erythrocytes (hRBCs) in a 'hot-cold' assay and caused cytotoxicity to 325 PMNs and MNLs. Both hemolytic and leukotoxic activity were found to be magnesium-dependent. RBC susceptibility to Beta-toxin correlated with the reported sphingomyelin content of each species. Scanning electron microscopy (SEM) demonstrated that 'hot-cold' incubation with Beta-toxin in the presence of magnesium caused significant morphological changes in the surface structure of both RBCs and PMNs. The changes included the formation of pits and membrane invaginations in the RBCs. The PMNs lost their ruffled membrane appearance and showed overall membrane disintegration. This study demonstrated that the viability of sphingomyelin-containing PMNs and MNLs was significantly decreased by the addition of Beta-toxin, indicating that this toxin does, in fact, have a leukotoxic nature. Leukocytes did not have significant membrane invaginations unlike toxin-treated RBCs; therefore, it is possible that leukotoxicity does not result from membrane lysis.
authors.affiliation: Department of Biological and Environmental Sciences, California University of Pennsylvania 15419, USA.
authors.forename: M J
authors.initials: MJ
authors.lastname: Marshall
authors.forename: G A
authors.initials: GA
authors.lastname: Bohach
authors.forename: D F
authors.initials: DF
authors.lastname: Boehm
mesh_terms: D001427:Bacterial Toxins / Q000008:administration & dosage / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004910:Erythrocyte Membrane / Q000187:drug effects; D006460:Hemolysin Proteins / Q000494:pharmacology; D006801:Humans; D007962:Leukocytes / Q000187:drug effects* / Q000648:ultrastructure; D008274:Magnesium / Q000494:pharmacology; D008855:Microscopy, Electron, Scanning; D013108:Sphingomyelin Phosphodiesterase; D013211:Staphylococcus aureus / Q000472:pathogenicity*
chemical_list: D001427:Bacterial Toxins; D006460:Hemolysin Proteins; D013108:Sphingomyelin Phosphodiesterase; C029878:hlb protein, Staphylococcus aureus; D008274:Magnesium