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.