The immunogenicity of K99 antigen in whole cell bacterins of Escherichia coli.

K99 antigen in Escherichia coli whole cell bacterins was immunogenic in rabbits and mice. Mice vaccinated subcutaneously and bled three weeks later had a serum antibody response which was dose dependent. The dose response on dilution of bacterins was shown to be mainly due to dilution of K99 antigen, rather than the reduction in adjuvant or bacterial cell concentration. The procedure appears to be a satisfactory one for immunogenicity testing of bacterins containing K99 antigen.

Organic acid proton donors decrease intestinal secretion caused by enterotoxins.

The effects of several weak acids on the secretory actions of cholera toxin and the heat-stable enterotoxin of Escherichia coli (ST) have been examined in ligated jejunal loops in weanling pigs. Ascorbic and acetic acids had no effect, but L-lactic acid reduced the net fluid secretion caused by cholera toxin. Glutaric acid and p-aminobenzoic acid blocked net fluid secretion caused by cholera toxin or by ST. Antisecretory effects were pH dependent for p-aminobenzoic acid in this study and for nicotinic acid in a previous report (6). At a pH of 5.0, p-aminobenzoic acid treatment increased lumen-to-blood sodium flux and decreased the blood-to-lumen sodium flux caused by cholera toxin. These weak acid effects were more marked on fluid fluxes in enterotoxin-treated loops than in control loops and persisted for 20-30 min after acid removal from loops. These findings are discussed in terms of requirements for antisecretory activity and possible modes of action of antisecretory compounds.

Nicotinic acid inhibits enterotoxin-induced jejunal secretion in the pig.

The use of nicotinic acid for preventing intestinal secretion caused by cholera toxin and by the heat-stable enterotoxin of Escherichia coli has been investigated in the weanling pig. Secretory effects were measured in ligated jejunal loops of halothane-anesthetized pigs by dilution of a nonabsorbable marker added to the loop fluid. Different routes of administration and different initial pH values for nicotinate solutions were studied to determine optimal conditions for secretory inhibition. The neutral sodium salt of nicotinic acid had no significant antisecretory activity under any conditions used in these trials. Inhibition of secretion was most effective with partly neutralized nicotinic acid at pH 4.5 added directly to loops containing enterotoxin. Net fluid secretion induced by cholera toxin or heat-stable enterotoxin of E. coli was prevented by this treatment. Reversal of secretion was not accompanied by any measurable changes in cyclic nucleotide concentration in intestinal mucosa. Nicotinic acid antagonism of a secretory step common to cholera toxin and heat-stable enterotoxin of E. coli but subsequent to cyclic nucleotide involvement is indicated by these data.

Isolation and partial characterization of two different heat-stable enterotoxins produced by bovine and porcine strains of enterotoxigenic Escherichia coli.

Heat-stable enterotoxins (ST-124 and ST-1261) have been isolated from two different enterotoxigenic Escherichia coli of bovine (124) and porcine (1261) origin. The enterotoxin preparations were isolated by ultrafiltration and ion-exchange chromatography and were both active in the suckling mouse test and pig ligated loop test in the nanogram range. The bovine (ST-124) enterotoxin was not stable to heating in its isolated form, and significant differences in amino acid composition were observed between the two enterotoxins. Although both toxins were active at similar levels in the suckling mouse and pig ligated loop tests, ST-124 lacked the ability to cause the profound secretory responses seen with ST-1261 in the weanling pig ligated loop.

Failure to reverse cholera toxin induced intestinal secretion by agents which decrease mucosal cAMP.

The feasibility of reducing intestinal secretion by the use of agents which decrease intestinal mucosal cAMP concentration has been investigated in the weanling pig and the rabbit. Three different agents for decreasing mucosal cAMP concentration were studied. The cyclic nucleotide phosphodiesterase activator, imidazole, significantly reduced mucosal cAMP concentrations only in the weanling pig. Intraluminal 2'-deoxyadenosine-3'AMP inhibited adenylate cyclase and caused a decrease in mucosal cAMP concentration in both the pig and the rabbit. The introduction of the heat-stable enterotoxin of Escherichia coli into pig jejunal segments also gave lowered mucosal cAMP concentrations. While these three agents effectively reduced cAMP concentrations in intestinal mucosa, they were ineffective in reducing the net fluid secretory effects of cholera toxin. Secretion caused by cholera toxin apparently persists independent of the temporary changes in cAMP concentration which can be induced by pharmacological agents.

Immunization of calves against enterotoxigenic colibacillosis by vaccinating dams with purified K99 antigen and whole cell bacterins.

Pregnant cattle were either vaccinated subcutaneously with (i) a suspension of purified Escherichia coli K99 pili, (ii) a Formalin-killed whole cell bacterin containing enterotoxigenic E. coli strain B44 (O9:K30;K99:H-), or (iii) a bacterin containing six different strains of bovine enterotoxigenic E. coli (multiple-strain bacterin), or were left as nonvaccinated controls. After birth, calves were allowed to nurse their dams and, at 12 to 14 h of age, were challenged orally with 10(11) cells of enterotoxigenic E. coli strain B44. Colostral antibody titers were determined against K99, K30, and O9 antigens of B44. In the nonvaccinated control group, 9 of 10 calves developed diarrhea and died within 24 to 72 h. Similarly, all six calves in the multiple-strain bacterin group developed diarrhea and four died. In contrast to calves in the two groups mentioned above, calves nursing cows vaccinated with either purified K99 or the homologous whole cell bacterin were protected against fatal diarrhea. There was a highly significant correlation (P less than 0.0005) between protection against fatal diarrhea and K99, but not K30 or O9 colostral antibody titers. Vaccination of cows with either purified pili or whole cell preparations containing sufficient K99 antigen may provide a means of preventing enterotoxigenic colibacillosis in calves.

Evidence for two heat-stable enterotoxins produced by enterotoxigenic Escherichia coli.

Heat-stable enterotoxins from a bovine and porcine strain of Escherichia coli were isolated and showed significant differences in amino acid composition and heat stability.