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Twenty-one adult volunteers were immunized at monthly intervals with three doses of purified type 1 M protein of group A Streptococcus. The soluble vaccine in buffer was administered by aerosol spray into the nares and oropharynx; 23 control subjects received a buffer placebo in the same manner. Antibody responses were observed in sera and nasal washings of some but not all vaccines. Approximately 30 days after the last dose, all subjects were challenged with homologus streptococci applied by swab to the phayngeal-tonsillar areas. In a double-blind system of evaluation, physical signs and symptoms were followed for assessment of infection. Illness was defined on the basis of a positive throat culture, fever, a twofold increase in white blood cell count over baseline, exudative pharyngitis, and cervical adenopathy. By these criteria four vaccinees and 11 controls were obviously ill. One vaccinee and six controls were questionably ill, fulfilling some but not all of the criteria. sixteen vaccinees and six controls were not ill (P less than 0.001). Positive throat cultures were observed in five vaccines and 19 controls (P less than 0.001). Penicillin was administered five days after challenge. No poststreptoccal sequelae or other complication were observed. Thus local immunization with M protein apparently can prevent both colonization and clinical illness after challenge with homologous streptococci.

In examining 170 samples of blood sera of healthy and sick individuals it was shown that with the aid of a simple and express antilipoproteinase test it was possible not only to reveal, but also to determine the titre of specific antibodies to the lipoproteinases of various serological M-types (2, 4, 22/12, 22 and 49) of a hemolytic streptoccus, group A. The antibodies to the enzymes were frequently revealed in the sera of adults and older children. The results obtained were still inadequate for the epidemiological or clinico-immunological analysis, but pointed to the principal possibility of using the antilipoproteinase test for the mentioned purpose.

Streptococci are amont the most common bacterial pathogens physicians encounter in practice. Infections with streptococci continue to occur with significant frequency despite the general sensitivity of these organisms to a variety of widely used antibiotics. In newborn infants and other special patient groups, streptococci may produce fulminant and fatal sepsis (Table 1). In normal children and adults, infections usually are short term and often mild or unrecognized but with the possibility of resulting, unpredictably, in nonsuppurative complications some weeks or months later. Although scarlet fever has become an unusual and clinically attenuated disease, its rashless analog, streptococcal pharyngitis, presents thorny problems in the differential diagnosis of symptomatic patients and in the detection of subclinical infections. Erysipelas now is a rare disease, but recent studies have confirmed that streptococci often are the primary etiologic agent in impetigo, another type of skin infection--with peculiar bacteriologic and epidemiologic features. Infections with group D streptococci have always been a special case because of their frequent resistance to penicillin, and group B streptococci (also somewhat resistant) present special problems in the perinatal period. Streptococci may appear in unexpected places or guises (see Table 1). Thus, the modern physician has little reason to relax in his vigilance for and knowledge of streptococcal infections.

Using the long chain test, and in some cases the bactericidal test, to measure antibody, the development of the immune response in horses to Str. equi has been followed. Long chain indices in excess of 5.0, accompanied by strong bactericidal capacity, were recorded in serums after the full 3-dose immunisation course with a commercial vaccine. The full course elicited the most satisfactory antibody titres declined within the 12 month post-vaccination period, thus providing support for the recommendation that yearly booster doses should be administered. The immune response in horses during 2 strangles outbreaks was compared with the response following vaccination. Recovered animals showed declining antibody levels 9 weeks after infection, and the evidence does not support longevity of the immune response after natural infection. Reaction to vaccination was observed in those animals with high initial antibody titre.

The results of complex surveillance of Streptococcus pyogenes, based on mass infections in the Czechoslovak army over a period of 11 years, are submitted. The epidemiological method revealed an incidence of cycles, lasting several years, of predominant pairs of T types, most of which could not be typed by their M protein. Auxiliary immunological surveys allowed the biological M activity of widespread strains to be estimated and probably form a basis for considerations on a negative long-term epidemiological type prognosis. The preliminary results presented offer prospects for a realistic methodological approach to the question of the type content of a future antistreptococcal vaccine.

Prospective studies of streptococcal pyoderma were carried out among black children enrolled in Project Headstart centers in Holmes County, Miss. Sera collected from 28 of these children in early October were tested for opsonic antibodies to one of two prevalent skin strains of group A streptococci isolated from them on one or more occasions over the preceding 3 months. The two streptococcal strains (A and B) belong to M-types previously unrecognized. Ten subjects (36%) had antibody to their homologous serotypes detectable by the indirect bactericidal test: this included 6 of 10 subjects infected with strain B but only 4 of 18 infected with strain A (P < 0.05). Of 17 children who had strains A or B isolated from skin lesions only, 12% developed type-specific antibodies (TSA) against the infecting serotype. In contrast, 11 subjects had these strains isolated from throat cultures (either with or without associated pyoderma), and 72% had detectable TSA (P < 0.01). There was no demonstrable relationship between the development of antibodies to streptococcal extracellular products or to non-type-specific cellular antigens and the development of TSA. These results demonstrate that type-specific immune responses do occur following infection with pyoderma streptococci. The frequency with which such antibodies develop is variable and appears related to a number of factors, including the immunologic properties of the infecting strain and the site of bacterial colonization. Pharyngeal carriage may represent an important mechanism for development of acquired immunity to skin strains of group A streptococci.

Antibodies against the group polysaccharide of group A streptococci were estimated by means of a haemagglutination reaction. In this reaction human erythrocytes of blood group O were sensitized with polysaccharide esterified with myristoylchloride. The optimal conditions of the reactions were determined by varying the ester group content in the antigen and the amount of ester used for sensitization. The specificity of the reaction could be established by reacting sensitized erythrocytes with homologous and heterologous sera and by absorption experiments. Antistreptococcal group A polysaccharide titres (ASPAT) and antibody levels to streptolysine O and DNase-B were compared in a group of 52 children with proved streptococcal infection and in 52 age- and season-matched controls. Antibody levels were significantly higher in the patient group than in the controls. In the ASPAT there was clearly less overlap between patients and controls than in both other reactions. In the patient group the ASO titres were raised above normal in 27 cases (51.9%), anti-DNase-B titres in 18 (34.6%), and ASPAT in 40 (76.9%). Taken together the three reactions gave a positive score in 51 cases (98.1%) in the patient group against 17 cases (32.7%) in the controls. A positive antibody response is usually defined as a rise of two dilution increments between the acute and convalescent sera. According to this definition the ASPAT showed a response in 42%, ASO and/or DNase-B in 42%, and the three reactions taken together in 68% of paired sera from patients. It is believed the ASPAT will prove a welcome addition to the diagnostic outfit when the presence of streptococcal infection in children is considered.

The lack of impressive quantitative differences in antibody to various streptococcal extracellular and cellular antigens among patients with acute rheumatic fever, acute glomerulonephritis, and following uncomplicated streptococcal infection has prompted investigation of qualitative aspects of the antibody response among these patients. By using a radiolabeled antigenically univalent hapten derived from streptococcal A carbohydrate, affinity of serum antibody to A-carbohydrate (A-antibody) was studied by an ammonium sulfate precipitation technique. The data obtained demonstrate average association constants (K0S) of acute rheumatic fever patient sera to be significantly lower than those of acute glomerulonephritis or streptococcal infection patients (P<0.02 and P<0.001, respectively). Further analysis of the data from hapten binding studies documents the fact that the radioimmune precipitin assay for the determination of A-antibody level is little influenced by K0 but directly correlates with the concentration of antibody binding sites. These data suggest that qualitative differences in A-antibody are present between rheumatic and non-rheumatic individuals. It is unclear whether the finding of low-affinity A-antibody among acute rheumatic fever patients reflects a generalized phenomenon or one restricted to the A-antibody-A-carbohydrate system.

Intravenous administration of live microorganisms to rabitts with cardiac catheters produces an experimental model of infective endocarditis. Despite the development of infected valvular vegetations, positive blood cultures, splenomegaly, and focal embolic renallesions, glomerulonephritis has not been found in these animals. In the present study, acute diffuse proliferative glomerulonephritis, featuring endothelial and mesangial proliferation, capillary occlusion, and leukocytic infiltration was produced in rabbits immunized withthe infecting agent prior to the establishment of left sided alpha-streptococcal endocarditis. Controls receiving immunization alone, immunization and sterileendocarditis, or infective endocarditis alone did not develop diffuse glomerulonephritis. Electron microscopic findings of occasional subendothelial electron-dense deposits and immunofluorescence deposition of IgG and C3 in a peripheral granular capillary pattern were consistent with an immune complex type nephritis. Decreased serum complement levels were demonstrated in those animals developing diffuse glomerulonephritis, and some animals developed circulating rheumatoid factor. In view of the morphologic findings and the necessity of preimmunization for development of glomerular changes, it is concluded that immune mechanisms play a role in the diffuse glomerulonephritis associated with this model of infective endocarditis.

Thirty-five children between 6 and 17 years treated in the clinic for scarlet fever, rheumatic fever and other non-streptococcal infections as controls, were tested by lymphocyte transformation to four streptococcal antigens. In all cases of scarlet fever and especially of rheumatic fever the lymphocytes were better stimulated by streptococcal products than in the control group. The SO and the MAP fraction showed a good stimulating activity. The response in the rheumatic fever patients was not influenced by the steroid treatment, nor by the stage of the illness. A parallelism with high humoral and cellular responses to SO at the beginning of the acute rheumatic fever was observed, followed by a dissociation of both responses during the evolution with the maintenance of the cellular one and the decrease of the ASO titre.

The state of cell-mediated immunity was measured by the morphological method of lymphocyte transformation with and without PHA, in a group of 56 patients with bacterial infections and in a group of healthy controls. The patients were divided into three subgroups according to the aetiology: 1. Patients with Gram-positive infections, 2. Patients with Gram-negative infections, and 3. mixed infections. The transformation values with PHA were significantly (t-test) higher in the Gram-positive subgroups than in the control group. Transformation in cultures stimulated by PHA was much higher in the subgroup of patients with Gram-positive aetiology; the difference between the values in the Gram-negative and in the Gram-positive subgroups was also significant. A decrease of transformation value below 50% was observed in 6 out of 18 patients with Gram-negative aetiology, whereas in the Gram-positive group it was noted in only 2 out of 21 patients.

Previous studies have shown the efficacy of parenteral immunization of volunteers with purified type 1 M protein against challenge with homologous streptococcui (J. clin, Invest. 52: 1885, 1973). A double-blind study was conducted on 21 adults immunized by aerosol-spray into the nasopharynx, and 23 controls who received saline placebo. Two booster doses were given at monthly intervals and approximately 30 days later vaccinees and controls were challenged with homologous streptococci (10-6/ml) by swabbing the pharyngeal-tonsillar areas. Throat cultures, leukocyte counts, temperatures and physical signs and symptoms were followed to assess infection. Illness was defined as a positive throat culture, oral temperature of larger than or equal to 38 degrees C, a WBC count of twice baseline or greater than 10,000 per mm-3, exudative pharyngitis and adenopathy. Of the 43 subjects, 13 were ill by all criteria (10 controls, 3 vaccinees [p smaller than .02]); 21 were well by all criteria (6 controls and 15 vaccinees); and 10 exhibited some but not all positive criteria (7 controls, 3 vaccinees). Positive throat cultures following challenge were obtained in 19 controls and 5 vaccinees (p smaller than 0.001). There was no correlation between the pre-challenge serum antibody titer and the development of subsequent illness. It is concluded that local topical immunization with a M protein vaccine offers significant type-specific protection against challenge with streptococci.

An outbreak of skin sepsis due to beta-haemolytic streptococcus group A M33, T3/13/B3264 occurred in a closed community. Staphylococcus aureus was also present in many of the lesions, but different strains were recognizable by phage typing. Environmental contamination was found. The outbreak was controlled by isolating infected boys and decontaminating the environment. Various aspects of the epidemiology and the pathogenesis of streptococcal infections are discussed.

The effect of oral and parenteral penicillin on the development of cellular and humoral immune responses in chimpanzees infected with group A streptococcal M-types 1, 5 and 12 was investigated. The interrelationship between type-specific antibody response and enhancement of phagocytic competence of polymorphonuclear neutrophils was documented. Penicillin depressed or suppressed type-specific antibody response depending on the mode and dose of administration, probably because of its effect on the streptococci responsible for antibody stimulation. Penicillin was not demonstrated to have a direct effect on phagocytic ability in vitro. Therefore the primary effect of antibiotic therapy is the indirect relationship to suppression or inhibition of type-specific antibody response to M-protein which results in a diminution of phagocytic competence.

The streptococcal infection cases from two outbreaks were serologically examined against two components of the streptococcal cellular wall; the M associated protein (MAP), by a latex agglutination test and the group A carbohydrate (A-CHO), by passive hemagglutination technique. Many positive cases with high levels of both antibodies were found, in rheumatic fever and glomerulo-nephritis, comparatively with the acute streptococcal infections. The differences were statistically significant. The results were correlated with the ASO titres and with the dermic cellular response.

An analysis of the cell surface antigens of Streptococcus equi is presented. Following extraction from cell walls, whole cells and hyaluronidase treated cells the immunoprecipitation patterns of the intact and protease treated components were developed with rabbit anti serum against S. equi. It was determined by mouse protection tests that the specificity of protective antibodies was directed toward a trypsin labile antigen. The presence of antigens similar to those designated M and R in Group A streptococci were found in S. equi.

A large number of bacterial species are causative agents of respiratory tract disease. The discussion will center on three infections selected because they represent different problems in control on the basis of epidemiologic and immunochemical factors. Group A hemolytic streptococci are common upper respiratory tract pathogens which may initiate severe non-suppurative sequelae such as rheumatic fever and glomerulonephritis. Recent progress concerning M protein vaccines will be reviewed. Pneumococci are still the most frequent cause of pneumonia at all ages. Pneumococcal vaccines are the prototype for purified polysaccharide vaccines since their effectiveness was demonstrated 30 years ago. The major problem in vaccination is the very large number of capsular serotypes. Pseudomonas aeruginosa represents the relatively new problem of gram-negative bacterial infections in the immunodepressed host. Demonstration of seven immunotypes as the cause of 90% of human infections has led to preparation of a multivalent vaccine composed of lipopolysaccharide antigens. Current knowledge of this vaccine will be discussed.

Streptozyme is a new serologic assay which is able to evidentiate 5 antistreptococcal antibodies at the same time (TAS, ASHA, ASK, DNase, NADase). This test has been carried out on 355 samples together with TAS, ASK and ASHA methods in order to value this method comparatively. The results showed that Streptozyme is a poorly reliable technique, looking at the false-positive or negative results obtained, but principally looking at the phenomena of no good reproducibility. We feel that at present it cannot be used in substitution of the classic serologic methods.