An enzymatic mutant of Shiga-like toxin II variant is a vaccine candidate for edema disease of swine.
Edema disease (ED) of weanling pigs is caused by an infection with Escherichia coli that produces Shiga-like toxin II variant (SLT-IIv). Pathology identical to that caused by ED can be duplicated in pigs that are injected with less than 10 ng of purified SLT-IIv per kg of body weight. Therefore, SLT-IIv was mutated to create an immunoreactive form of the toxin that was significantly reduced in enzymatic activity. Initially, purified SLT-IIv was treated with formaldehyde which abrogated cytotoxic activity. Pigs were vaccinated with the toxoid (100 micrograms) to determine whether a toxoid was a viable vaccine candidate and whether young pigs were capable of mounting an immune response. Although the pigs developed a neutralizing antibody titer (1:128 to 1:512) 28 days postinjection, they also lost weight and developed ED lesions. The deleterious effect of the toxoid appeared to result from residual enzymatic activity or a reversion to a toxic form. An alternative method, site-directed mutagenesis, was employed to consistently reduce the enzymatic activity of SLT-IIv. Glutamate at position 167 of the mature A subunit was replaced by aspartate (E167D), and arginine at position 170 was replaced by lysine (R170K). These mutations reduced cytotoxic activity 10(4)-fold and 10-fold, respectively, while the enzymatic activities were decreased 400-fold and 5-fold, respectively. The activity of a toxin that contained both mutations (SLT-IIvE167D/R170K) closely resembled that of SLT-IIvE167D. When position 167 was replaced by glutamine (E167Q), the cytotoxic activity decreased 10(6)-fold and the enzymatic activity decreased approximately 1,500-fold. Pigs that were vaccinated with purified, mutant toxin designated SLT-IIvE167Q developed a neutralizing antibody titer of 1:512 21 days postinjection, and their tissues were free of ED lesions. These data suggest that SLT-IIvE167Q may represent an effective vaccine against ED.
Gordon
V M
VM
Department of Microbiology, Uniformed Services University of the Health Sciences, Bethesda, Maryland 22908.
Whipp
S C
SC
Moon
H W
HW
O'Brien
A D
AD
Samuel
J E
JE
D000818:Animals; D000831:Animals, Newborn; D001427:Bacterial Toxins / Q000276:immunology*; D001428:Bacterial Vaccines / Q000276:immunology*; D004488:Edema Disease of Swine / Q000517:prevention & control*; D004768:Enterotoxins / Q000276:immunology*; D009154:Mutation; D022641:Shiga Toxin 2; D013329:Structure-Activity Relationship; D013552:Swine; D014121:Toxoids / Q000276:immunology
D001427:Bacterial Toxins; D001428:Bacterial Vaccines; D004768:Enterotoxins; D022641:Shiga Toxin 2; D014121:Toxoids