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Escherichia coli P16 was shown to produce two heat-stable toxins '(ST) with differing biological activity. The toxins were separated by methanol extraction, and the first, STa, was methanol soluble, partially heat stable, active in neonatal piglets (1 to 3 days old) and infant mice, but inactive in weaned pigs (7 to 9 weeks old); the second, STb, was methanol insoluble, active in weaned pigs and rabbit ligated loops, but inactive in infant mice. It is therefore suggested that use of suckling mice as indicators of ST production will fail to identify certain STproducing strains.

Summary Grass pollen extract was reacted with glutaraldehyde, and used in the form of a lyrosine adsorbate for the treatment of hay fever. The incidence of systemic reactions was reduced by the chemical treatment to negligible levels, permitting the therapeutic‐schedule to be reduced to three doses without impairment of clinical effectiveness. The levels of pollen‐specific IgG antibody induced by the chemically modified material were similar to those induced by pollen‐tyrosine or by alum‐precipitated pyridine extract. The increase in IgE antibody induced by glutaraldehyde‐pollen‐tyrosine was lower than that induced by pollen‐tyrosine: only the falter achieved significance.

While studying the involvement of cyclic adenosine 3',5 '-monophosphate (cAMP) in the fluid secretion caused by heat-stable enterotoxin (ST) from Escherichia coli P16 in infant mice, it was noted that the culture filtrate containing ST also contained large amounts of cAMP. The present paper details attempts to obtain a cAMP-free ST preparation. The organisms were grown in a defined medium, and the heated culture filtrate was concentrated by reverse osmosis. After methanol extraction of the filtrate, which removed 80% of the nonactive solids, the methanol-soluble ST was further purified by gel filtration through a Sephadex G-10 column. The first fraction recovered after gel chromatography contained ST with a negligible amount of cAMP. Treatment with methanol did not adversely affect the enterotoxic activity. Certain parameters of the infant mouse model have been investigated, and using our ST preparation it has been found that animals remain responsive up to 15 days of age with an optimum assay time of 2 h after toxin challenge.

The ability of antisera to lipid A, induced in rabbits by immunization with lipid A complexed to various carriers, to protect mice against gram-negative infection and to inhibit the fluid loss caused by an enteropathogenic strain of Escherichia coli in the piglet ligated gut was investigated. No significant protection was obtained in either case, although passive hemolysis and quantitative precipitation tests showed the presence of antilipid A antibodies in the sera. Fluorescent antibody studies suggest that the lipid A is in a cryptic position on the surface of smooth strains of gram-negative bacteria.

Partially purified heat-stable enterotoxin obtained from Escherichia coli strain F11/P155 caused an accumulation of cyclic GMP in the intestines of 8-day-old mice.

S T R A I N S o f Escherichiu coli enteropathogenic for swine have been shown to secrete enterotoxins in vitro (Smith and Halls, 19673;Gyles and Barnum, 1969). These enterotoxins cause accumulation of fluid when they are introduced into ligated intestinal loops prepared in rabbits or pigs (Smith and Halls, 1967~). The strains of E. coli involved are believed to produce two different types of enterotoxin, namely a heat-labile non-dialysable immunogenic toxin (LT) and a heat-stable dialysable toxin (ST).The use of infant rabbits for the detection of LT has been reported by Kutas, Vetesi and Semjen (1974). The present paper described the response of infant rabbits to LT in greater detail, and the evaluation of them as small-anima1 models for the detection of LT.The quantity of LT required for assessment of the infant-rabbit model necessitated a large-scale fermentation, and during the course of this work, it was reported that fermentation of strains of E. coli in the presence of lincomycin (Levner, Wiener and Rubin, 1977) or mitomycin C (Isaacson and Moon, 1975) increased LT synthesis. Hence, it was decided to investigate the effect of these agents on the yield of LT from E. coli strain P307, and these results are also reported here. The preliminary assessment of increase in yield of LT was determined in rabbit ligated-intestinal loops. MATERIALS AND METHODSOrganisms. E. coli strain P307 (08: K87: K88a, b) was obtained from Dr H. Williams Smith (Houghton Poultry Research Station, Houghton, Huntingdonshire). It produced LT and ST. Five substrains resistant to lincomycin 300 pg/ml were obtained by sequential transfer of liquid cultures into media containing increasing concentrations of lincomycin, followed by purification of resistant clones as reported by Levner et al. (1977). The Lin' phenotype was stable in the absence of the drug.Culture media. Large-scale growth of strain P307 was produced on syncase medium (Finkelstein, Norris and Dutta, 1964) modified by the substitution of glucose for sucrose and by the addition of yeast extract 0.6%(w/v).The lincomycin-resistant strains were grown on the modified casamino acids and yeast extract medium described by Evans, Evans and Gorbach (19736) in the presence and absence of linomycin 250 pg/ml.

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