Comparative in vitro leukotoxin production of three bovine strains of Fusobacterium necrophorum.
The in vitro leukotoxic activity of 3 bovine isolates of Fusobacterium necrophorum which varied in pathogenicity were compared. Monolayers of mouse peritoneal macrophages were exposed to culture filtrates from each F necrophorum strain, and cell viability was determined, using the trypan blue dye exclusion test. Two methods were used for production of the leukotoxin: (1) medium M-1 continuous dialysis sac cultures and (2) brain-heart infusion agar plate cultures. Supernatant cultural fluids containing the leukotoxin were subjected to membrane-partition chromatography, using ultrafilters with approximate molecular weight (mol wt) exclusion limits of 100,000, 10,000, 2,000, and 500. All ultrafiltrates had a cytotoxic effect on the monolayers. Cytotoxic activity was not found in the ultrafilter residues or in the control media ultrafiltrates. Comparative study of leukotoxin production indicated that F necrophorum 2101, type A, produced the most leukotoxin; F necrophorum 2030, type AB, produced slightly less leukotoxin; and F necrophorum 2035, type B, produced small amounts of leukotoxin. Endotoxin activity, as demonstrated by the mouse lethality test, was found in the residues of the XM-100A ultrafilter (100,000 mol wt), but not in the filtrates. Culture supernatant fluids and the XM-100A ultrafiltrates were positive for endotoxin, using the limulus amebocyte lysate assay; however, the other ultrafiltrates with lower mol wt exclusion limits were negative.
D000818:Animals; D002417:Cattle; D004731:Endotoxins / Q000302:isolation & purification; D005098:Exotoxins / Q000096:biosynthesis* / Q000302:isolation & purification; D005675:Fusobacterium necrophorum / Q000378:metabolism* / Q000472:pathogenicity; D008264:Macrophages / Q000187:drug effects; D051379:Mice; D014774:Virulence
D004731:Endotoxins; D005098:Exotoxins; C018579:leukotoxin