Five strains of encapsulated group A streptococci of different serological types, each with a glossy and a matt variant, were studied to compare the rôles of the M substance and the hyaluronic acid capsule in virulence of these microorganisms. The results indicated that both contribute to the virulence of group A streptococci but that the M antigen is the more fundamental factor. Encapsulated variants, both glossy and matt, were slightly less susceptible to phagocytosis than those from which the capsule had been removed with hyaluronidase. Glossy variants, containing no M substance, were readily phagocyted; matt, M-containing variants were resistant to phagocytosis except in the presence of anti-M serum when they became fully susceptible. Only the M-containing, matt strains were mouse-virulent. Mice were protected against infections with these strains: (a) By removal of the capsule with hyaluronidase, which resulted in slight protection, but only against 10 M.L.D. Early and intensive treatment was required to produce this effect; i.e., simultaneous injection of enzyme and streptococci followed by prolonged enzyme therapy. (b) By a single injection of anti-M serum administered the day before inoculation of the streptococci, which resulted in protection against 100,000 M.L.D. (c) By combined use of enzyme and anti-M serum, an additive effect of the two protective agents occurred, which resulted in protection against 1,000,000 M.L.D.

A study of collagen was initiated several years ago in relation to work on the pathogenesis of rheumatic fever since it is generally believed that the primary injury in this disease is to collagen, the ground substance, or both (1-5). The chemistry and structure of collagen have been studied extensively (6--19) but little is known of its biological properties. 1 The studies to be reported below relate primarily to the antigenicity of collagen. Review of Previous WorkNageotte (20,21) showed that when the tail tendons of the rat are placed in dilute acetic acid (1:25,000 dilution in distilled water) the collagen fibers slowly swell and over a period of time go into solution. He also demonstrated that fibers reconstituted from this acid solution by the addition of sodium chloride retain the tinctorial properties of native collagen. Wyckoff and Corey (22) demonstrated that the x-ray diffraction pattern of this reconstituted material was like that of the native tendon. Schmitt, Hall, and Jakus (9) and others (23, 24) have found that fibers reformed from dilute acid solutions either by neutralization or the addition of salt retain the characteristic striations of native collagen as observed in electron micrographs.It is generally agreed that gelatin, a soluble derivative of collagen, is non-antigenic (25, 26). Loiseleur and Urbain (27) reported complement-fixing antibodies in the sera of rabbits immunized with an acid solution of rat tail tendons prepared as described by Nageotte. Hopps (28) demonstrated antibodies to catgut and sheep intestine; however, there were cross-reactions with sheep serum. With highly purified preparations of sheep collagen obtained from Dr. F. O. Schmitt he reports agglutination with anticatgut rabbit serum in a dilution of 1:80; however, another sample of the highest

Although the presence in convalescent human sera of type-specific antibodies to group A streptococci has been reported by several workers (1-10), the specificity has not always been supported by unequivocal evidence. The newer knowledge of the different types of group A streptococci provided by Lancefield (11,12) and Griffith (13), and of the antigenic structure by the former, has made possible a new approach to the problem.A reliable and sensitive method of determining type-specific circulating antibodies would help greatly in solving the question of type-specific immunity; it would also provide a technique for investigating epidemiological and clinical problems resulting from streptococcal infections and their sequelae. Typespecific antibodies in the blood of patients recovering from group A streptococcal infections have been investigated by a variety of methods: agglutination, precipitation I complement fixation, mouse protection, opsonic, and bacteriostatic techniques. Both slide and macroscopic agglutination reactions have proved unsatisfactory because of non-type-specific reactions and also because of the difficulty in obtaining uniformly stable bacterial suspensions. Precipitin and complement fixation tests with human sera and M extracts reveal many cross-reactions with extracts of heterologous types. The mouse protection test requires strains of high mouse virulence which are infrequent in streptococci freshly isolated from human sources; this test apparently requires a higher antibody content than that usually found in convalescent human sera; moreover, large amounts of sera are necessary. The opsonic index technique is frequently difficult to interpret. The ba~teriostatic test, although dependent upon the opsonization by the serum and phagocytosis of the streptococcal cell byleukocytes, has yielded reliable and easily interpreted information concernLug the presence of type-specific antibodies. In conformity with previous investigators, the term bacteriostasis used in this report implies not merely the inhibition of bacterial growth but also the destruction of microorganisms by sensitization with convalescent serum and their subsequent phagocytosis by leukocytes.

No abstract

During the course of routine typing of group A streptococci recovered from nasopharyngeal cultures of patients suffering from hemolytic streptococcal infections of the upper respiratory tract, it was noted that, although there was no change in the colony morphology on sheep or rabbit blood agar plates, the streptococci obtained during convalescence often produced less type-specific M substance than the streptococci isolated during the acute phase of infection. It was also observed that the streptococci isolated in the convalescent period were often susceptible to the bacteriostatic action of normal children's blood, whereas the microorganisms isolated in the early period of the infection were invariably resistant.These findings suggested that following infection in man the antigenic composition of the hemolytic streptococcus may undergo changes similar to those known to occur in the pneumococcus (1) and diphtheria bacillus (2) following natural infections with these bacteria.Todd (3) and more recently Ward and Lyons (4) have shown that the ability of hemolytic streptococci to multiply in normal human blood is a reliable and sensitive method for distinguishing different strain variants and for determining mouse virulence. Hare (5) has also reported that saprophytic strains of hemolytic streptococci isolated from the human birth canal can be differentiated from pathogenic strains by their susceptibility to the bactericidal action of human blood. Furthermore, various workers (6-9) have shown that the typespecific M antigen is necessary for the exhibition of virulence of group A streptococci; and that the presence of this protein antigen characterizes the matt variant, whereas the glossy variant produces little or no M substance.Since previous reports of hemolytic streptococcal variation have been confined for the most part to in vitro or animal studies, a similar study was undertaken to investigate this phenomenon during the natural course of infection inman. For this purpose the cultures were obtained at weekly intervals from patients during the acute, convalescent, and carrier stages of their streptococcal respiratory infections; and the individual-strains were subjected to tests for their ability to resist the bacteriostatic action of normal human blood and to synthesize the type-specific M protein antigen.