The Immunizing Antigens of Erysipelothrix rhusiopathiae. The Role of the L-Antigen

Abstract

SUMMARY : Vaccines were prepared by forniolizing cultures of Erysipelothrix rhusiopathiae and precipitating with aluminium hydroxide gel (AG). Fifty per cent of mice were protected against about a thousand lethal doses. Two (Ru and VYS) of eleven strains yielded highly potent vaccines.Serum in the medium increased the potency of vaccines and for one of the strains liver extract was needed in the medium to give potent vaccines. In general, vaccines protected well against heterologous strains but protection was relatively poor against one of these.A soluble antigen present in culture fluids induced in rabbits antibodies protecting mice and agglutinating suspensions of E. rhusiopathiae grown in the presence of serum. This soluble antigen was thermo-labile. In these respects it resembles the L-antigen, differing only in being detached from the surface of cells. Soluble antigen was precipitated with AG, yielding fairly good vaccines. Cells freed from media were also precipitated, yielding rather better vaccines. It is suggested that cells yield vaccines of high potency when they have produced and retained a relatively large amount of L-antigen.Alert hiolate-killed cells of Erysipdothrix rhusiopathiae grown in serum-containing media are capable of inducing in rabbits circulating antibodies protective for mice (Gledhill, 1947). Preliminary experiments with pigs indicated that whereas some circulating antibodies were also produced by intravenous injection of killed cells, a stronger response was elicited by suspensions of formalin-killed cultures from which most of the cells had been removed by centrifugation (Gledhill, 1948). Traub (1947) showed that mice and pigs could be actively immunized with cultures precipitated with aluminium hydroxide and that both cells and a soIuble antigen present in cultures were necessary for producing fully active vaccines. Moreover, such active vaccines were only produced by a few strains of E . rhusiopathim.However, Traub did not give the technical details of his methods. The purpose of this investigation is (1) to confirm that killed vaccines precipitated by aluminium hydroxide gel immunize mice effectively; ( 2 ) to confirm the existence of Traub's soluble antigen; (3) to determine its relationship to the L-antigen developed on the surface of cells grown in serum-containing media (Gledhill, 1947). M E T 1 10D SMaintenance of strains. I n general, strains were maintained on serum agar slopes and subcultured a t monthly intervals. Good immunizing strains were lyophilized. Strains for challenging mice were passaged through normal mice a t frequent intervals and cultures made on serum agar slopes were kept a t 72-2

Isolation and Characterization of a Protective Antigen-Containing Particle from Culture Supernatant Fluids of Erysipelothrix rhusiopathiae

“…However, treatment with trypsin resulted in the loss of all activity, suggesting the presence of an essential protein or peptide moiety in the protective antigen. In addition, muramidase, which acts upon B-1, 4-glucosidic linkages in glycosamino To compare further the protective antigen present in the 20P fraction to that which has been previously reported (7), the heat stability of the protective activity of the 20P fraction was studied. Samples (20 ml), containing 8 mg of the 20P fraction in 0.05 M sodium phosphate buffer (pH 6.8) were subjected to 1 hr of exposure in water baths at 50, 64, 75, 85, and 95 C. Results of protection tests (Fig.…”

“…Samples (20 ml), containing 8 mg of the 20P fraction in 0.05 M sodium phosphate buffer (pH 6.8) were subjected to 1 hr of exposure in water baths at 50, 64, 75, 85, and 95 C. Results of protection tests (Fig. 3) indicated that there was no loss of activity at 50 C. At 64 C, the point at which Gledhill (7) reported that all activity was lost, there was still about 25% of the protective activity present. The protective activity remained at this level through treatment at 85 C but at 95 C again decreased.…”

“…In 1947, Traub (16) reported that certain strains of E. rhusiopathiae grown in the presence of horse serum were capable of producing enough protective antigen in vitro to be useful as killed vaccines. Later studies (4) indicated that these strains, which belonged to Dedie's serotype B, were protective against both serotypes A and B. Traub's finding (16) that part of the protective activity of these vaccine cultures was the result of soluble protective antigen dissociated from the bacterial cell was confirmed by Gledhill (7).…”

“…Nani (12) found that, although most of the protective activity of whole cells was extracted in the protein fraction when using the Boivin method, a minor portion of the activity was associated with a glycolipid fraction. The protective antigen is destroyed by heating to 64 C (7), is sensitive to freezing (11), and is resistant to ethanol at room temperature (7). An investigation was undertaken to characterize the protective antigen of E. rhusiopathiae further.…”