Protective Effect of Immunization with Heat-Labile Enterotoxin in Gnotobiotic Rats Monocontaminated with Enterotoxigenic
            <i>Escherichia coli</i>

The respective contributions to protection of the route and dosage of primary and booster immunizations with Escherichia coli heat-labile enterotoxin were evaluated in rats. The degree of protection was determined by challenge with toxin and viable bacteria in ligated ileal loops, and the serum antitoxin response was assayed by enzyme-linked immunosorbent assay. Primary immunization was effective only when given by the parenteral route. The degree of protection was enhanced by a fivefold dosage increase in the primary parenteral immunization in rats given constant dosages of booster immunizations either parenterally or perorally, but not by further dosage increases. In contrast, the degree of protection rose when dosages of the booster immunizations were increased over a 25-fold range. Four weekly peroral, but only two biweekly parenteral, booster immunizations were necessary to achieve strong protection; biweekly combined parenteral and peroral booster immunizations yielded both strong, immediate and extended protection. The degree of protection against the toxin correlated with that against viable bacteria and with elevated serum antitoxin titers: all seven groups with a protection index of >5 against the toxin had strong protection against heat-labile toxin-producing strains and fourfold or greater increases in the antitoxin titers, whereas none of the nine groups with a protection index of <3 was protected against bacteria or had an equivalent antitoxin response. These observations show that once an adequate parenteral primary immunization is given, the degree of protection is influenced primarily by the dosage of the booster immunizations, the necessary number of which is dependent on their route of administration. Acute diarrheal disease caused by intestinal colonization with enterotoxigenic strains of

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Respective contributions to protection of primary and booster immunization with Escherichia coli heat-labile enterotoxin in rats

Klipstein¹,

Engert²

1981

“…The degree of protection was less in some instances when the toxins were given exclusively by the parenteral route, and the B subunit failed to provide any protection whatsoever against the LT'/ST' strain when given by this approach. It seems probable that the protection against this strain that is induced by immunization with LT toxin forms is directed against the LT produced by this strain (22). We suspect that the lesser degree of protection that was observed in some instances in animals immunized exclusively by the parenteral route was due to the fact that this approach stimulates principally systemic IgG antibodies, whereas the combined parenteral-PO immunization regimen provides protection derived from both systemic antibodies and IgA antibodies produced locally within the intestinal mucosa (25,32,34,35).…”

Section: Discussionmentioning

confidence: 99%

“…The first demonstrated that in rats immunized with various different regimens and dosages of PM LT, those which had a PI > 5 against challenge with the toxin also consistently had >50% reduced secretion when challenged by the LT+/STstrain (21). The second study showed that immunization with those dosages which yield these values in ligated loops also provides protection against the secretory effect of intestinal colonization with either an LT+/STor an LT'/ST' strain in gnotobiotic rats (22).…”

Section: Methodsmentioning

confidence: 97%

Protective Effect of Immunization of Rats with Holotoxin or B Subunit of Escherichia coli Heat-Labile Enterotoxin

Klipstein

Engert

1981

Self Cite

The relative immunogenicities of three forms of the Escherichia coli heatlabile enterotoxin (LT), the holotoxin, its B subunit, and the polymyxin-release form (PM LT) were compared by immunizing rats with various dosages of each given exclusively by the parenteral (IP/IP) or peroral (PO/PO) routes or by a combination of the two (IP/PO). The degree of protection was evaluated by challenge in ligated ileal loops, and the serum antitoxin response was determined by an enzyme-linked immunosorbent assay with homologous antigens. When given by the PO/PO route, each LT antigen provided only weak protection against the toxin and virtually none against viable LT-producing strains; serum antitoxin titers were not significantly increased. When the toxins were given after a parental primary immunization by either the IP/IP or the IP/PO routes, each LT antigen provided a dose-related increase in serum antitoxin titers and in the degree of protection against the toxin as well as against viable strains which produce LT alone (LT'/ST-) or in combination with the heat-stable toxin (LT'/ ST'). The degree of protection against viable bacteria, particularly the LT'/ST' strain, was stronger in animals which received booster immunizations by the PO route. When expressed on the basis of molar equivalents, holotoxin provided significant protection (a protection index of >5 against toxin challenge and >50% reduced secretion with bacterial challenge) with 4 to 15 times fewer moles than PM LT and up to 50 times fewer moles than the B subunit. These observations indicate that, on the basis of molar equivalents, the holotoxin (which contains one A plus five or six B subunits) is a more potent immunogen than either PM LT (which contains one A and probably one B subunit) or the B subunit.

“…The results of the present study establish the effectiveness of immunization with this vaccine in a second experimental animal model, rabbits. Protection in both animal models was demonstrated by means of challenge in ligated ileal loops; the applicability of this technique to protection under challenge conditions in which the entire intact intestine is colonized by LT-producing enterotoxigenic strains of E. coli has been confirmed in gnotobiotic rats immunized with LT (20).…”

Section: Discussionmentioning

confidence: 92%

Protection in rabbits immunized with a vaccine of Escherichia coli heat-stable toxin cross-linked to the heat-labile toxin B subunit

Klipstein¹,

Engert²,

Houghten³

1983

Rabbits and rats were immunized with a vaccine consisting of synthetically produced Escherichia coli heat-stable toxin cross-linked by the carbodiimide reaction to the B subunit of biologically produced porcine heat-labile toxin. The vaccine contained 50% of each toxin component by weight and antigenicity; the toxicity of the heat-stable enterotoxin component was reduced by >600-fold. Two or three peroral immunizations with vaccine containing 1,000 antigen units of each component raised greater-than-threefold increases in specific mucosal immunoglobulin A antitoxin titers to each component in all animal groups. Protection index values for challenge with either heat-labile or heat-stable toxins in ligated ileal loops were 3.4 to 4.0 in rats immunized by a parenteral primary immunization followed by two peroral booster immunizations, >9 in rabbits immunized by these routes, and >8 in rabbits given just three peroral immunizations. The antigenicity of the B-subunit component of the peroral vaccine was protected equally well against gastric acidity either by pretreatment with cimetidine or by delivery of the vaccine encapsulated in pH-dependent microspheres. The vaccine did not cause diarrhea when given perorally to any of the experimental animals or evoke fluid secretion when instilled into rabbit ligated ileal loops. These observations (i) confirm the effectiveness of this vaccine as an immunogen in a second animal model, (ii) establish that it is effective when given exclusively by the peroral route, and (iii) provide further evidence regarding its lack of toxicity. Enterotoxigenic strains of Escherichia coli cause acute diarrhea by means of elaborating heat-labile or heat-stable enterotoxin (LT or ST, respectively), either singly or together. LT and its B subunit are immunogenic (13,16); lowmolecular-weight, haptenic ST becomes immunogenic when coupled to a high-molecularweight carrier (7,9). Immunization of experimental animals with either of these toxins raises a specific antitoxin response that provides protection against viable strains of heterologous serotypes which produce that particular toxin. Thus, immunization with either LT or its B subunit provides protection only against LTproducing strains (LT'/ST-) (13,16), immunization with ST yields protection only against STproducing strains (LT-/ST+) (14,18), and antitoxin to each toxin form contributes to protection against strains which produce both toxins together (LT'/ST+) (13, 17a). Conjugation of ST to LT yields an immunogen that provides protection against strains which produce either form of toxin since ST acquires immunogenicity as a result of its coupling to LT and the antigenicities of both toxin components are largely re-tained although their toxicities are greatly reduced (15).We recently described a vaccine for enterotoxigenic E. coli based on cross-linking synthetically produced ST to the nontoxic B subunit of LT (17). Since the immunogenicities of ST and the B subunit are approximately the same on a weight basis, the optimal composition of this ...