Demonstration of the Association of Specifically Different Alpha Streptococci with Various Diseases and Methods for the Preparation and Use of Specific Antiserums and Vaccines in Diagnosis and Treatment

Despite the existence of a great volume of evidence that certain strains of alpha or green-producing streptococci are causative of widely different diseases, much confusion still exists. This is true of both ill-defined³³ and clinically well-defined epidemic⁴² and nonepidemic⁴¹ diseases. The reasons for confusion are many, yet quite obvious. Clinicians often do not fully appreciate experimental evidence as regards proof of etiologic importance of inciting agents and bacteriologists all too often work with inadequate methods. Streptococci belonging to this group that have specific properties are usually highly sensitive to oxygen and require special mediums for their isolation and for maintenance of specificity. Conventional mediums now in general use do not suffice. Corroboration of the observations of my colleagues and me has or has not been obtained, depending on whether the principles embodied in our methods^4,5,8,11,13 have or have not¹⁵ been used. Alpha streptococci having specific properties are associated with diseases at present attributed to virus as now understood. The criterion of death or survival of inoculated animals as evidence of virulence does not suffice in studies on the etiologic importance of this group of organisms. Careful search for lesions, and cultures from lesions if found, in inoculated animals that die as well as in those that survive, are essential to prove that the organisms have specific action and etiologic importance. The principle of dissociation or mutation^16,30–which makes difficult their classification by cultural and serologic means and the preparation of really specific vaccines– is not considered sufficiently or is overlooked entirely by different investigators. What has been said regarding causation applies with even greater force to the whole question of passive and active immunization with antiserums and vaccines.

By the use of special methods which have been developed during extended studies on the etiologic importance of these streptococci, not only have we been able to isolate them consistently from atria of infection and from systemic lesions in different diseases, but we often have succeeded in reproducing or simulating, in important respects, the corresponding disease pictures. This has been accomplished in animals by various methods of inoculation of strains freshly isolated in dextrose-brain broth–by intravenous and intracerebral and other methods of inoculation, by feeding and, most important, by inducing chronic foci of infection through the devitalization of the teeth of dogs and inoculation of the respective streptococci into them.³⁴ Other workers have had similar results.^8,11,13 Moreover, it has been shown that the respective streptococci which localize electively in certain tissues or organs produce “endotoxin” and “exotoxin” which cause lesions in the very same tissues as those in which lesions are produced by the corresponding living streptococci. This was demonstrated by the injection separately of the washed organisms killed by heat or formalin and the filtrate of actively growing cultures. These basic observations indicate that general and localized manifestations of disease may be the result of hypersensitivity or allergy specifically related to the streptococci and their products which have specific affinity for certain tissues.

In addition to the consistent isolation of streptococci in the reproduction of the respective lesions in animals, further evidence in favor of their etiologic importance has been obtained by other wholly unrelated methods. By a special staining method²⁷ diplococci, resembling those in dextrose-brain broth cultures of the streptococcus from poliomyelitis, have been found with the light microscope (fig. 1) and in unstained films with the electron microscope, infiltrates of encephalitic and poliomyelitis viruses. The streptococci from different diseases on isolation in dextrose-brain broth were found to have distribution curves of cataphoretic velocity characteristic not only of different and of closely related diseases but, most important of all, of certain streptococcal diseases involving tissues derived from the mesoderm or ectoderm and entoderm, respectively, or from both (fig. 2). The distribution curves of cataphoretic velocity of the streptococci and dextrose-brain broth were similar in the different diseases studied to those of the washed streptococci in distilled water directly from atria of infection. The need for the study of these streptococci on isolation in dextrose-brain broth before specific properties were lost, and the importance of mutation or dissociation were illustrated especially well by these cataphoretic studies. It was found that on exposure of streptococci in this medium to the high frequency field of electric energy for varying time intervals the distribution curves of cataphoretic velocity characteristic of different diseases could be induced in any given strain and that concomitantly changes in localizing power characteristic of the distribution curves of cataphoretic velocity often occurred. As the distribution curve of “neurotropic” streptococci became “arthrotropic” the streptococci lost “neurotropic” virulence and acquired “arthotropic” virulence and vice versa.

Horses have been immunized in parallel manner by the repeated intravenous and subcutaneous injection of increasing doses of different strains of streptococci freshly isolated in dextrose-brain broth. The number and interval of injections, the doses of the different strains, and the bleedings were made in parallel manner. The only variable was the disease from which the different strains were isolated.

The strains used for immunization were isolated either from the systemic lesions of the patient or from respective lesions produced in animals by appropriate inoculations of streptococci isolated from atria of infection. Pure cultures of the streptococcus then were grown for eighteen hours in large amounts of 0.2 per cent dextrose broth. The organisms were harvested with a continuous-feed centrifuge and immediately suspended in glycerin (two parts) and 25 per cent solution of sodium chloride (one part), the growth from approximately 500 cc. of broth being added to 1 cc of menstruum. The suspensions then were stored in the refrigerator and appropriate dilutions in solution of sodium chloride were made as needed for injection into horses.

To obtain highly specific anti-streptococcic serums it does not suffice to inject streptococci after prolonged cultivation nor as grown on artificial mediums during the long period of immunization, because of their tendency to undergo change.^19,23 The results obtained in animals, in cataphoretic studies on streptococci and on the diagnostic value of anti serums together with a review on elective localization, bibliography and interpretation of the studies on focal infection are available.^6,34

The Precipitation Reaction

For precipitation tests swabbings were made from the nasopharynx high up behind the palate with cotton-wrapped aluminum wire swabs bent to a suitable angle. Care was taken not to touch the tongue with the swab. The adherent material was washed off in 2 cc. of gelatin (0.2 per cent) Locke solution by thorough shaking and cleared by centrifugation. The serum used as antigen was obtained from the blood of patients drawn before breakfast. The cleared extracts and clear serums, respectively, were superimposed on the respective anti serums with as little admixture as possible in small precipitation tubes. Readings were made, after incubation for one and a half hours at 35° C. and after refrigeration overnight, under the edge of the shade of a 75 watt electric light in a dark room against a non-reflecting black velvet cloth background. The degree of precipitation was usually slight but only those reactions were recorded as positive in which there was no question of clouding at the interphase.

The precipitation reactions between extracts of nasopharyngeal swabings of persons ill with various diseases and the homologous and heterologous anti-streptococcic serums are summarized in table 1, and those obtained with the serums from patients and these anti-streptococcic serums are summarized in table 2. A high incidence of reactions and a high degree of specificity with the homologous serums were obtained. The serum of horses during the acute stage of encephalomyelitis⁴⁴ was precipitated as specifically as the serum of persons having encephalitis by the encephalitis anti-streptococcic serum prepared with strains from the disease in human subjects. The incidence of reactions with heterologous antiserums was greater with extracts of nasopharyngeal swappings, as should be expected, than with the serums, and in both groups of diseases was highest in those serums prepared with streptococci most nearly resembling the specific strains, such as with the epilepsy serum in the group of patients having epidemic or postoperative persistent hiccup or ether convulsions.⁴⁶ Proper dilutions of cleared extracts of the different streptococci preserved in dense suspension in glycerin and salt solution often were precipitated specifically by the homologous antiserums.

The Reaction After Cutaneous Injection of Antibody

It was found in previous studies²⁹ that intradermal injection of the whole anti-streptococcic serums prepared by intravenous injection exclusively, and in the present study that injection of the euglobulin fraction of the serum of horses immunized by both intravenous and subcutaneous injection, were followed immediately (five to ten minutes) by an erythematous-edematous reaction at the site of injection of the homologous antibody,^32,36,39 but by little or no reaction at the sight of injection of heterologous antibody. This reaction is similar to the reaction first described by Foshay⁷ in tularemia and by Tamura⁴⁸ in venereal lymphogranuloma. In our test if the patient’s skin or serum contains antigen immunologically related to the antigen or streptococcus with which the reacting antibody is prepared, an immediate transient flare occurs (fig. 3). The flare is taken to indicate the presence of unneutralized antigen in the serum or skin due to an infection by streptococci antigenically identical or related to the streptococcus used to prepare the reacting antibody.

In order to determine more precisely than nature of this reaction, identical numbers of the dead streptococci from poliomyelitis and arthritis were injected intracerebrally into two series of rabbits and Macacus rhesus monkeys. The cutaneous tests in rabbits proved unsatisfactory. the monkeys receiving the poliomyelitic streptococci reacted positively to intradermal injection of the poliomyelitic euglobulin but not to the arthritic euglobulin. Those receiving the arthritic streptococci reacted to intradermal injection of the arthritic euglobulin but not to the poliomyelitic euglobulin. Precipitation reactions with the serums of both monkeys and rabbits and the respective homologous antiserums were positive during the period when the cutaneous reaction was positive. Moreover, the poliomyelitic streptococci remained in the spinal fluid much longer than the arthritic streptococci but did not appear in the knee joint fluid (fig. 4), whereas the arthritic streptococci disappeared promptly from the spinal fluid but appeared, often in easily demonstrable numbers, in the knee joint fluids, as well Illustrated in figure 5. Control punctures of spinal canal and joints before injection proved that spinal fluid and knee joint fluids were sterile and free from bacteria, and all cultures of suspensions injected and of test spinal fluids and joint fluids proved sterile. All organisms, the skin reactivity to homologous euglobulin and the precipitation reaction disappeared in forty-eight to seventy-two hours after injection. This experiment was repeated with a different set of suspensions of the respective streptococci with the same results.

Striking evidence of specificity of the cutaneous reaction to injection of antibody prepared with streptococci isolated from persons having the corresponding disease was obtained in nearly all of a large number of patients tested. The results are summarized in table 3, in which both the average reaction and the percentage incidence of reactions 5 sq. cm. or more are shown.

Routinely, 0.03 cc. of a 0 per cent solution, in saline containing 0.2 per cent phenol, of the euglobulin fraction (obtained by diluting the serum 1 to 10 in slightly acidulated distilled water) of the different antiserums, and of normal horse serum diluted 1 to 10, was injected as superficially as possible into the skin, usually of the forearm. The size of the maximal flare of erythema was outlined with pen and ink and then traced in transparent paper. From such tracing the area of erythema was determined. Maximal reaction usually occurred within five to ten minutes, after which the erythema rapidly disappeared. Slight late (three to four or more days) reaction associated with itching sometimes occurred owing to local sensitization to horse protein.

The cutaneous reactions to the different homologous antibodies were so nearly alike in persons having diseases manifesting similar symptoms and affecting tissues or organs of the same embryologic origin that these are considered in groups for the sake of brevity.

The degree of reaction was highest after injection of the encephalitis euglobulin in the group of patients having encephalitis or spasmodic torticollis. The group having persistent hiccup, epilepsy or other convulsions, in which spasms of muscles suggesting involvement of the brain were the outstanding manifestations, reacted maximally to the encephalitis and epilepsy euglobulins. The persons who had acute polioencephalitis and the group of persons who had chronic poliomyelitis, progressive muscular atrophy, or multiple or amyotrophic lateral sclerosis, in accord with the involvement of brain and cord, reacted maximally to the encephalitic and poliomyelitic euglobulins. The persons who had epidemic poliomyelitis in which only the spinal cord was affected reacted specifically, chiefly to the poliomyelitic euglobulin.

The groups that had chronic infectious arthritis, fibrositis, myositis or heart disease reacted maximally and very specifically to the euglobulin prepared with streptococci from arthritis and rheumatic fever. The group representing infections of the respiratory tract–influenza, colds, bronchitis, asthmatic bronchitis and bronchopneumonia–reacted similarly and maximally to the globulin from the serum of horses immunized with streptococci from influenza.

The group of persons suffering from gastrointestinal diseases–ulcer of the stomach, chronic ulcerative colitis and epidemic gastroenteritis–reacted maximally to the euglobulin prepared with the streptococcus of ulcerative colitis. The twenty persons who had ulcer of the stomach or duodenum included in this group reacted even more strongly to the euglobulin prepared with the streptococcus from ulcer than to the ulcerative colitis euglobulin. The average reaction to the former and clinical cases of ulcer measured 13 sq.cm. and in 77 per cent of the cases there were reactions measuring 5 sq. cm. or more, whereas the reaction to the ulcerative colitis euglobulin approximated that for the gastrointestinal group. The reactions to the ulcer euglobulin in the groups of persons having encephalitis or arthritis were far less than those of patients having ulcer.

The reactions of well persons to the different euglobulins were uniformly slight, indicating that the beta streptococci normally present in well persons are different from those in persons having symptoms attributable to specific strains. The distribution curves of alpha streptococci from well persons remote from epidemic respiratory infections were like those of specific strains after prolonged cultivation on artificial mediums.²⁵ The former streptococci, as the latter, also lacked specific virulence.^17,34 The reactions to injection of normal horse serum diluted 1 to 10, as recorded in table 3, and to a 10 per cent solution of normal horse euglobulin in special instances were similar and uniformly slight, aside from those in a few persons sensitive to horse serum.

In extended studies on the diagnostic value of the cutaneous test, especially in connection with poliomyelitis and influenza, a striking difference in the reactivity of the skin to heterologous euglobulins was noted in certain groups of patients and well persons. This was found to be dependent, in part, on density of population in which persons were residing, whether in large cities or in rural communities. The reactions, as usual, to homologous euglobulins were maximal in both groups but reactions to heterologous euglobulins were greater in the urban than in the rural groups. This has been observed repeatedly and suggests subclinical but immunizing infection by antigenically different streptococci. This observation is in accord with the generally accepted view that city dwellers have a higher level of resistance when exposed to epidemic respiratory and other infections than persons residing in the country. A relatively higher incidence of reactors to the euglobulins characteristic of epidemics was found among well persons, whether exposed directly or not, within epidemic zones than among the persons remote from epidemics, irrespective of density of population. This is taken to indicate the wide prevalence of the positive streptococcus and perhaps the corresponding virus kind of subclinical immunizing infections, and–it would seem–explanation as to why epidemics, especially in rural communities, run their course usually within eight weeks, and why epidemics in which enduring immunity to the disease in question is conferred do not recur except at long intervals. Specific antibody content of convalescent serum was usually not sufficiently high to yield clear cutaneous reactions. A few serums from persons convalescing from acute epidemic poliomyelitis²⁹ and from recurring encephalomeningoradiculitis with fibromyositis³³ gave suggestive results.

Methods and Results of Agglutination Tests

The difficulties encountered in agglutination experiments with streptococci in general and with the green-producing group in particular, when made by conventional methods, have been largely overcome by the use of methods which we have developed. Green-producing streptococci which are causative of certain diseases have been found highly sensitive to oxygen. Ordinary mediums often do not suffice for their isolation, a fact not sufficiently considered in studies on etiology and in the preparation of vaccines. Hence, the primary isolations are made by us in the highly favorable medium, dextrose-brain broth,²² which affords a gradient of oxygen tension, a low oxidation-reduction potential, and other conditions highly favorable for growth of sensitive streptococci. The dextrose-brain broth consists of 0.2 per cent ordinary dextrose broth adjusted to pH 7.2, and the soft dextrose-brain agar also used likewise consists of dextrose broth containing 0.2 per cent agar, to both of which approximately one part of pieces of freshly obtained calf brain to six or seven parts of broth in tall (10 cm.) columns is added before autoclaving.

Pure cultures of freshly isolated strains were obtained not by plating on blood agar but by inoculation of animals or by making serial dilution cultures alternately in dextrose-brain broth and dextrose-brain agar at steps of 10 to 10,000, depending on the character of the inoculum.³¹ The streptococci to be used for agglutination experiments and for the preparation of prophylactic and therapeutic vaccines were grown usually for eight to eighteen hours in dextrose-brain broth. The supernatant culture was transferred to test tubes or bottles and centrifuged. The supernatant cleared broth was decanted and the streptococci were suspended routinely in the menstruum of glycerin-salt solution so that the suspensions contained approximately 100 times as many streptococci per cubic centimeter as the dextrose-brain broth culture. After thorough mixing the suspensions were placed in the refrigerator.

The dense suspensions in glycerin-salt solution and the respective antiserums were diluted in physiologic solution of sodium chloride containing 0.2 per cent phenol for the agglutination tests. The density of the suspensions was adjusted to approximately twice that of a dextrose-brain broth culture. The hyperimmune serums were diluted 1 to 5, 1 to 25, 1 to 125 and 1 to 625, and the serums of patients 1 to 10, 1 to 20, 1 to 40 and 1 to 80. Equal parts of the respective suspensions and serum dilutions were mixed and the setup was placed at 40 to 50°C. for eighteen hours, when readings were made, instead of after being kept at 37°C. for one or two hours and then in the refrigerator overnight, as is usually done. The degree of agglutination in the different dilutions was recorded according to the arbitrary scale of 0 to 4. In summarizing results agglutination was considered as specific in the serum that showed greater agglutination than occurred in any of the other antiserums used.

The results of agglutination tests made in this way with homologous and heterologous whole antistreptococcic serum or globulin concentrates are summarized in table 4. The results with strains from closely related diseases were so similar as to permit placing them in representative groups for brevity. The results in each of the different groups were highly specific, the incidence of agglutination being far higher in the homologous than in the heterologous antiserums. Although occasionally, as table 4 indicates, several tests were made on different cultures isolated from the same patient, these different cultures were as variable in their reactions as the total population of cultures isolated from the different patients, and the total results are not affected by this procedure. This was true also of the distribution and results of the several extracts and serums tested from the same persons in several of the groups summarized in tables 1 and 2. Therefore; the results are given in percentage of cultures, extracts or serums tested rather than percentages of cases.

In the agglutination tests, as was noted in the precipitation and cutaneous tests, the incidence of marked specific action of the epilepsy antiserum was limited to the strains of streptococci isolated in the cases of persistent hiccup and ether convulsions. In the case of streptococci isolated in studies of encephalitis, the high point of agglutination was by the antiserum prepared with streptococci isolated in studies of encephalitis in human beings. This serum also agglutinated specifically the strains of streptococci isolated in studies of equine encephalomyelitis. In the group of cases of acute poliomyelitis and progressive muscular atrophy maximal agglutination occurred in the poliomyelitis antistreptococcic serum. In the group of cases of multiple or amyotrophic lateral sclerosis in which both spinal cord and brain are affected, agglutination was high in both the poliomyelitis and the encephalitis antistreptococcic serums. The incidence of specific agglutination of the strains isolated in arthritis was by far the highest in the arthritis antistreptococcic serum; in those from chronic ulcerative colitis and epidemic gastroenteritis by the antiserum prepared with the diplostreptococcus; of ulcerative colitis, and in the group of strains from persons having colds, influenza, asthma or bronchitis by the antiserum prepared with the streptococcus from influenza.

The incidence of specific agglutination of streptococci isolated in twenty-three cases of myasthenia gravis, not recorded in table 4, was very high (74 per cent) in the antiserums prepared with the streptococcus isolated in studies of myasthenia gravis,^4,37,38 and next highest in the antiserum prepared with the streptococcus isolated in studies of arthritis. Absorption of the different antiserums with homologous streptococci removed their specific precipitating and agglutinating power. Streptococci from persons having miscellaneous diseases were not markedly nor differentially agglutinated by any of the antistreptococcus serums used.

The agglutinin content of the serum of persons having certain low grade illness is often too small to permit of clear-cut demonstration, even by our methods. However, well marked evidence was obtained in some instances of an increase of specific agglutinins during convalescence or continued infection. The results of experiments with a large number of strains and serums in acute and chronic encephalitis, poliomyelitis and arthritis are summarized in table 5. Well marked evidence of specific agglutinins in each of these diseases was found. Similarly, an increase in agglutinins specific for the strains of streptococci isolated consistently in the St. Louis epidemic of encephalitis occurred during convalescence. Thus, the incidence of agglutination, over the strains of streptococci isolated, in 170 tests made with eleven serums obtained one to seven days after illness began was 62 per cent; at eight to fourteen days with six serums it was 81 per cent of 132 tests, and at fifteen to twenty-one days with forty-five serums agglutination of the encephalitis streptococcus occurred in 84 per cent of 478 tests. In contrast, fifteen control serums used under identical conditions gave an incidence of agglutination of only 24 per cent of 228 tests. There was a similar, although a less marked, increase in agglutinating titer of these convalescent serums over the streptococci isolated in studies of poliomyelitis, but no increase of the agglutinating power over strains of streptococci isolated in arthritis and cholecystitis used as controls. Easily demonstrable increased agglutinating power over streptococci isolated from the blood during the acute stage of equine encephalomyelitis and from the brain after death developed in the serum of horses during convalescence from the disease.

Results From the Use of Antistreptococcic Serums and Streptococcic Vaccines

Results from the use of the antiserums prepared in the way indicated in the treatment of epidemic poliomyelitis^35,43 and of encephalitis^10,18 have been reported. Concomitantly with subjective improvement noted after approximately 0.5 cc. per kilogram of body weight of the globulin concentrate of the antiserum (one part euglobulin containing chiefly bacterial antibody and two parts pseudoglobulin containing chiefly antitoxic antibody) had been given, there occurred a prompt (two to four hours) diminution in the reactivity of the skin to reinjection of the respective euglobulins, but the reactivity reappeared to a lesser degree in eighteen to twenty-four hours. Two intramuscular injections per day for several days usually sufficed to bring the temperature and cutaneous reactivity to normal with concomitant alleviation of symptoms. Similar, as yet unpublished, results have been obtained with the encephalitis and epilepsy antistreptococcic serums in a series of cases of persistent epidemic or postoperative hiccup, in which the patients usually reacted strongly to both the encephalitis and epilepsy euglobulins. Similar striking results have been obtained in acute influenza with the influenza antistreptococcic serum. Improvement, sometimes striking, or staying of the process occurred in cases of chronic encephalitis,¹² chronic poliomyelitis, chronic ulcerative colitis³ or chronic infectious arthritis, by intramuscular injection twice weekly of subsensitizing doses of the respective concentrated antiserums. The schedule of dosage in these cases consisted of the intramuscular injection of 0.1 cc. and increasing by 0.1 cc. up to two cc., provided untoward reactions did not occur and favorable effects were noted. Detailed reports of the further use of antistreptococcic serums and vaccines are beyond the scope of this paper.

Results, often striking, from the use of the vaccines prepared from the suspensions of streptococci in glycerin-salt solution in prevention and treatment of experimental infections in animals and in epidemic and other diseases in human beings and horses have been set forth in a series of reports.^{2,12,14,25,40,45} Vaccines prepared from streptococci that have been preserved in glycerin-salt solution are much less toxic and more promptly antigenic²⁵ than corresponding vaccines prepared with streptococci directly from cultures. By virtue of these properties they are better suited not only for prophylaxis but especially for treatment than when prepared directly from cultures. No untoward reactions⁹ have occurred after the injection of many thousands of properly gauged doses of the streptococcic vaccines prepared in this way.

Comment and Conclusions

Studies on alpha or green-producing streptococci associated with certain epidemic and nonepidemic diseases are reported. Special methods were found necessary for the consistent isolation of strains that had specific properties, for the demonstration of specific virulence, for the maintenance of specific antigenicity and for the preparation of highly specific antiserums and vaccines. Studies to be reported elsewhere indicate that the respective specificities, although clear-cut on isolation in each of the diseases studied, are related to environmental, seasonal and other influences.

Analysis of the data in tables 1, 2, 4 and 5 on the differences between the various serums from specifically immunized horses and normal horse serum, and the differences between the serums obtained or cultures isolated from persons suffering or convalescent from specific infections and similar serums or cultures from well persons, shows that these differences were statistically significant, except in a few cases in which the numbers were too small to make comparison possible. The striking parallelism of results obtained by the widely different methods used–inoculation of animals, electrophoresis and the precipitation, cutaneous, agglutination and agglutinin and precipitin absorption reactions–indicates especially their etiologic importance. The closely parallel results obtained from the use of the cutaneous and agglutination tests in the arthritic-rheumatic group of patients are in strict accord with the report by Baggenstoss and Rosenberg¹ of the finding of cardiac lesions identical with those of rheumatic fever in fourteen (56 per cent) of twenty-five cases of chronic infectious arthritis studied.

The mildness or even absence of symptoms suggestive of infection, the small number of apparent absence of organisms in systemic lesions, and the extreme sensitivity to specific vaccines, noted especially in chronic cases, suggest that a local or generalized allergy specifically related to the respective streptococci and to their toxic products is often responsible for the symptoms and progressive course in some of the diseases studied.

It is, of course, to be understood that the evidence, while quite conclusive in some diseases studied, is only suggestive in others. For example, the evidence indicating the possible importance of streptococci in the causation of epilepsy has been obtained in only a relatively small number of cases. Cultures of the blood at the time of seizure have yielded the streptococcus repeatedly. Intracerebral inoculation into animals of suitable doses of the streptococcus freshly isolated from blood and atria of infection produced as the outstanding manifestation localized and generalized spasms of muscles and sometimes recurring generalized convulsive seizures.²⁶ The cutaneous reaction was usually maximal with the homologous euglobulin and the precipitation reaction was consistently positive between the whole homologous antiserum and the serum of patients at the time of seizures. The streptococcus was agglutinated specifically by the homologous antiserum and the antibodies were removed specifically from the antiserums in absorption experiments with the streptococcus. Vaccines prepared with this streptococcus seemingly have produced cure in a number of cases of early epilepsy

The favorable results obtained in studies on the etiology and treatment by antistreptococcic serums and vaccines of chronic poliomyelitis, progressive muscular atrophy, multiple and amyotrophic lateral sclerosis, as well as of thrombophlebitis and pulmonary embolism^20,21 (not included in the tables), also are considered only as highly suggestive.

The nature of the causal relation of alpha streptococci having specific properties to the virus diseases studied and their viruses, as now understood, whether synergistic, dissociative, mutational or phasal, is not entirely clear. Our studies of encephalitis of the fox²⁴ and horse,⁴⁴ of poliomyelitis in man and monkey,²⁸ and of epidemic influenza in man indicate: (1) that the relation is probably chiefly phasal and that these diseases are primarily specific streptococcic infections; 2) that during the course of such infections a virus phase of the streptococcus develops; (3) that the respective streptococci and corresponding viruses are antigenically related, and (4) that with the respective streptococci it is possible to prepare preventive and curative vaccines and antiserums for these as for the other diseases studied.

The methods we have developed are relatively simple, and their general adoption, or the development of better ones in harmony with the underlying principles embodied, would result, it is believed, in a better understanding of the nature and treatment of the puzzling group of diseases associated with or caused by highly specific strains of alpha or green-producing streptococci.

 

References Cited: