The Use of Precipitin Analysis in Agar for the Study of Human Streptococcal Infections

Abstract

It is well known that streptococci can cause a variety of infections which are sometimes followed by non-invasive sequelae, e.g., rheumatic fever, chorea, etc.The factors involved in the production of these postinfection complications, and their mechanisms are not clearly understood. In view of the specificity of these sequelae relative to streptococcal disease, it seems likely that one or more particular products of the organisms are etiologically involved in some way. It has long been clear that a number of streptococcal substances are released in the tissues during the course of human infection, as evidenced by antibody rises at the time of convalescence (1). However, these bacterial products have only been discovered on a hit-or-miss basis until recently. The development of precipitin analysis in agar (2, 3) has furnished a tool for determining the total number of antibody responses following streptococcal infection in man, and thereby the total number of antigenic components released in vivo. Such studies carried out in this laboratory several years ago (4, 5) demonstrated that the numbers of streptococcal components which reach antibody-forming sites in significant amounts during human infections were surprisingly large. In addition, unexpectedly, most of the antigens thus detected were apparently of extracellular origin, and represented secretion products of the microorganisms. Very few antibody responses were found against the bacterial cellular extracts prepared in a variety of ways. Furthermore, pooled gamma globulin from normal human adults proved to be extraordinarily rich in these anti-streptococcal antibodies, thus indicating very frequent and widespread streptococcal disease in the population at large. Confirmatory results have been obtained by other groups (e.g., references 6, 7).Since it is of primary importance to clarify the identity and possible pathological significance of these streptococcal factors thus shown to be secreted in vivo in humans, an extended program was begun to purify and characterize these antigens. Early re-* These investigations were assisted by research grants from the

Postgenomic Analysis of Four Novel Antigens of Group A Streptococcus : Growth Phase-Dependent Gene Transcription and Human Serologic Response

“…For more than 40 years, it has been known that patients with GAS infections produce antibodies to a large number of extracellular proteins (16)(17)(18)(19)(20)24), but most of these antigens have not been identified. Availability of genome sequences has permitted virulence factors and therapeutic agent candidates to be identified very rapidly by comparative genomics and other postgenomic methods (2,3,14,21,24,32,35,37).…”

An Analysis of the Toxic Actions of Purified Streptolysin O on the Isolated Heart and Separate Cardiac Tissues of the Guinea Pig

“…~ These preparations were made according to methods previously described (8)(9)(10)(11)(12) and had been used in other studies (4)(5)(6). The Group A toxin consisted entirely of protein and had a titer of 160,000 hemolytic units (HU) per milligram of dry weight; the Group C material, which was 67% protein, had a tlter of 40,000 HU/mg.…”

“…Antiloxin.--Normal pooled human gamma globulin (E. R. Squibb & Sons, New York), a 16% solution containing known streptococcal antibodies with an anti-streptolysin O titer of 2000 Todd anti-streptolysin units/ml (8)(9)(10)(11), was furnished through the American Red Cross. Two purified plasma proteins, human serum albumin (Cutter Laboratories, Berkeley, Calif.), and bovine gamma globulin (Armour & Co., Chicago, Ill.), were used for control of antitoxin.…”

The Analysis of Streptococcal Infections

“…Studies of this sort with human streptococcal infections have been carried out during recent years in this laboratory (3)(4)(5)(6)(7)(8). It has been found that sera from patients with rheumatic fever are very rich in precipitating antibodies to extracellular streptococcal products, but contain few if any to cellular extracts prepared from the same strain.…”

“…When large scale purifications of crude culture supernates containing these antigens were carried out by means of continuous flow electrophoresis combined with column chromatography on calcium phosphate gels, it was then found that the systems were even more complex (5,7). At least sixteen extracellular antigens were detected for which humans had spontaneously developed antibodies.…”