title: Phospholipase A(2) in rabbit tears: a host defense against Staphylococcus aureus.
abstract: OBJECTIVE
  This study analyzed rabbit tears for anti-staphylococcal activity, the role of phospholipase A(2) (PLA2) in this reaction, and the ability of enzyme inhibitors to promote bacterial survival.
  
  
  METHODS
  Contact lenses with Staphylococcus aureus were applied to scarified rabbit eyes. The colony-forming units (CFU) per cornea or lens were determined and pathology was scored by slit-lamp examination (SLE). The bactericidal activity was measured by incubating bacteria with rabbit tears or PLA2 at 33 degrees or 37 degrees C. Radiolabeled S. aureus was incubated with PLA2 or tears to quantify the release of a membrane component that was identified by thin-layer chromatography. Inhibitors of these reactions were also analyzed.
  
  
  RESULTS
  Application of Staphylococcus, on contact lenses, to rabbit corneas resulted in bacterial killing and limited inflammation. Incubation of tears and bacteria (1:1; v/v) in tryptic soy broth at 33 degrees C decreased CFU approximately 4 logs. Tears (> or =30 microl) or PLA2 (> or =30 U) incubated with bacteria in phosphate-buffered saline were bactericidal. PLA2 (> or =0.2 U) or tears (> or =2 microl) cleaved bacterial membranes, liberating arachidonic acid. Spermidine or tetracaine inhibited cleavage of bacterial membranes by tears or PLA2 and spermidine promoted bacterial survival and growth in tears. Tears (60 microl) killed >99% of the bacterial inoculum, whereas bacteria incubated in tears plus spermidine approximately doubled in number.
  
  
  CONCLUSIONS
  PLA2 in rabbit tears kills Staphylococcus by hydrolyzing bacterial membranes to release arachidonic acid. Spermidine and tetracaine inhibited PLA2 activity and spermidine protected Staphylococcus from PLA2 in rabbit tears.
authors.affiliation: . Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans 70112, USA.
authors.forename: J M
authors.initials: JM
authors.lastname: Moreau
authors.forename: D O
authors.initials: DO
authors.lastname: Girgis
authors.forename: E B
authors.initials: EB
authors.lastname: Hume
authors.forename: J J
authors.initials: JJ
authors.lastname: Dajcs
authors.forename: M S
authors.initials: MS
authors.lastname: Austin
authors.forename: R J
authors.initials: RJ
authors.lastname: O'Callaghan
mesh_terms: D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D002855:Chromatography, Thin Layer; D015169:Colony Count, Microbial; D003261:Contact Lenses / Q000382:microbiology; D003315:Cornea / Q000382:microbiology*; D015818:Eye Infections, Bacterial / Q000382:microbiology*; D007634:Keratitis / Q000382:microbiology*; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000502:physiology*; D054467:Phospholipases A2; D011817:Rabbits; D013095:Spermidine / Q000494:pharmacology; D013203:Staphylococcal Infections / Q000382:microbiology*; D013211:Staphylococcus aureus / Q000254:growth & development* / Q000302:isolation & purification; D013666:Tears / Q000201:enzymology*; D013748:Tetracaine / Q000494:pharmacology
chemical_list: D013748:Tetracaine; D016718:Arachidonic Acid; D010741:Phospholipases A; D054467:Phospholipases A2; D013095:Spermidine