Partial purification and characterization of the NAG vibrio enterotoxin

Abstract

The culture filtrates of non-cholera vibrios (NAG vibrios) which possessed toxic activity were concentrated, precipitated with (NH4)2SO4 and fractionated on a column of Sephadex G-100. Three fractions were obtained. The first fraction contained mainly lipopolysaccharide which possessed no enterotoxic activity measured by the fluid accumulation in the ligated rabbit ileal loop as well as no activity in the skin permeability test. However, it was toxic for mice after i.v. administration (LD50: 600 microgram per mouse). The second fraction contained one antigenic protein and revealed four bands in the region of albumin migration distance in polyacrylamide electrophoresis. The size of the molecules from the second fraction was estimated by the elution volume from the G-100 Sephadex and polyacrylamide electrophoresis at approximately 50,000-70,000 daltons. This fraction had marked proteolytic activity. The third fraction contained proteins. In biological experiments only the second fraction possessed the enterotoxic activity identical to that of the original filtrate. The coexistence of the permeability and hemorrhagic factor in this material was proven by means of the rabbit skin test and a fast toxic effect for infant mice after intragastric inoculation. The enterotoxic activity characteristic of cholera enterotoxin was demonstrated in the ligated ileal loop of rabbit only after administration of 5-10 fold concentrated culture filtrate and the second fraction.