Abstract
In the first chapter of the paper a review is made of the literature concerning the immunology of the more important deep mycosis. Types of antigens, immunological reactions used, cross reactions observed and the results supplied by the immunological study are described. In this way, a review of the literature about Coccidioidomycosis, Histoplasmosis, North American Blastomycosis and South American Blastomycosis is made. Regarding the South American Blastomycosis, suggestions are made to achieve a better knowledge on the immunology of this disease. In chapter two, the material and methods used in complement fixation and precipitin reactions are described. In this chapter, some findings about the antigen are included. First, a table (table I) with data regarding 22 batches of antigen prepared up to 1959 are presented. The data refer to the strains of Paracoccidioides brasiliensis, the days of culturing and the optimal fixing capacity of the antigens with 3 and 6 units (50% hemolysis) of complement. An experiment on preservation of the antigen is recorded. It can be kept in the ice-box under sterile conditions without preservative for a period of time as long as three years. Under these conditions there is no modification in the properties of the antigen. In this chapter it is also reported an experiment about immunization of rabbits using as antigen the polysaccharide and a suspension of yeast-like cells of P. brasiliensie. It was not possible to obtain any antibody in the experiments which were carried out with only two rabbits, and the results have been included in order to call the attention about the difficulty of obtaining hyperimmune serum from these animals. Another experiment recorded in this chapter deals with the possibility of obtaining "polysaccharide antigen"
from each strain employed in the preparation of the antigen (most times 8 strains were used - table I). In order to make this study, iso-fixation curves were prepared as advised by Almeida in 1956. The protocols of these experiments are given in tables II to XIV and the results are represented in diagrams 1 to 3. Each one of the eight antigens was obtained from a separate strain of P. brasiliensis and it was also tested in the precipitin reactions with a human serum from a patient with South American Blastomycosis. All antigens gave positive results as can be seen in table XV. In this chapter, the principal features of the complement fixation reactions which had been studied in a previous work as well as the technique followed in the precipitin reaction are also mentioned. In chapter III, the results of precipitin and complement fixation reactions in 220 patients of South American Blastomycosis are presented, each serum was tested by the two kinds of reaction at the same time. The results of complement fixation (Wadsworth, Maltaner & Maltaner) and of the precipitin reactions in 22 cases of localized forms of South American Blastomycosis are presented. Some cases had had the disease for less than one year (table XVI) and others for more than one year (table XVII) when their blood was collected. Tables XVIII and XIX show the results observed in the disseminated forms of South American Blastomycosis in 59 patients with less than one year of duration and 107 patients with more than one year of duration before their sera were tested. In table XX, results concerning 32 patients considered clinically cured when their blood was collected are presented. In table XXI, there are the results of the immunological follow-up of 10 patients with the localized form of South American Blastomycosis who were chosen according to increasing periods of illness duration before collection of the first blood specimen. It can be seen that some of these cases were classified in this group at the beginning of the observation but according to their immunological behavior they were cases of disseminated forms of South American Blastomycosis (e.g. case A.A. in the table XXI). In table XXII, we have the immunological follow-up of fifteen cases of disseminated forms of the disease chosen according to increasing periods of illness duration before collection of the first blood specimen. In table XXIII, likewise, we have the immunological follow up of seven cases of clinically-cured forms. Some cases in the last three tables had only one serum tested and they were included on account of the period of evolution of the disease and in order to demonstrate the general behavior in each group. In this chapter are also presented the results of the immunological study of 1 case of South American Blastomycosis associated with leprosy, 2 cases associated with tuberculosis, 1 case with Addison's disease, because of the metastatic localization of the illness in the adrenals, 5 cases with metastatic localization of the disease in the central nervous system, 2 cases in which the diagnosis of cancer was made after the immunological examination suggested that they were not cases of South American Blastomycosis and two cases of South American Blastomycosis in which the diagnosis was first made by the immunological data. Finally, we have the results of an immunological study among 47 relatives of the patients which was made with precipitin and complement fixation reactions in order to show the inter-human transmission (table XXVIII). Chapter IV records the discussion of the results obtained regarding the antigen employed, the preservation of the antigen and the preparation of antigen from different strains of Paracoccidioides brasiliensis. Regarding the patients of South American Blastomycosis, an attempt is made to adjust the results to a very general scheme of the evolution of an infection when a microorganism penetrates the human body. When this happens, the following may occur: a) infection without disease; b) mild infection - generally localized forms; c) severe illness - disseminated forms. It is emphasized that the author had not a good antigen for making skin tests and that the first of these forms, e.g., blastomycosis infection without disease, has not been demonstrated, Studies made in 1959 by Prof. Lacaz et alii suggest that this form exists in the South American Blastomycosis like in other deep mycosis. Regarding the localized forms of South American Blastomycosis it may be remarked: a) the rarity with which they were observed in this research, only ten per cent; b) the time of appearance of the two kinds of antibody; c) the low titer of the complement fixing antibody; d) the persistence of the antibody in the sera of the patients after treatment. The precipitin was the first antibody to appear in the blood of the patients and the first to disappear after treatment, Diagram 5 records the immunological evolution of a typical case of this form of South American Blastomycosis. Regarding the disseminated forms of South American Blastomycosis, the higher titer observed in the complement-fixation reaction may be remarked, These titers may come down with some rapidity after treatment in cases supposed to be of acute disseminated forms, as shown in diagram 6. In other cases, treated with sulphadrugs without improvement, the titer remains high as shown in the diagram 7, where the titer only carne down when the patient was subjected to other treatments like sulpha and D2 vitamin in December, 1957, and Amphotericin B in June, 1958. In this case, clinical and serological relapses could be seen. In other chronical cases, a clinical cure may be achieved without decrease in the titer of fixing antibodies (as we can see in Table XXII - case J. R.). In diagram 8 there is a case of serological relapse during the year of 1957 without symptoms or signs of clinical relapse. Regarding the precipitin in the disseminated forms, it is present when the disease is in activity, The follow-up of the clinically cured cases shows that the complement-fixing antibody remains in the sera after the treatment, and after precipitin has vanished, as revealed by the high percentage of complement-fixation reaction positive in these cases. A discussion is made of the cases associated with other illnesses, of the cases of metastatic localization in adrenals and central nervous system, and of the investigation in relatives of the patients. Chapter V records the conclusions of the present work. First, about the antigen used (a polysaccharide f'rom Paracoccidioides brasiliensis): a) this antigen is good for the precipitin and complement-fixation reactions; b) it can be obtained easily using various strains of Paracoccidioides brasilensis; c) it can be titrated very well by block titration; d) it can be kept in the refrigerator for a period as long as 3 years without any modification in its properties; e) the yield of polysaccharide is quantitatively different in different strains of P. brasiliensis. The immunological study of 220 patients of South American Blastomycosis allows the following conclusions:
Using the two kinds of test - precipitin and complement fixation reactions - circulating antibodies may be demonstrated in 98.4% of the patients of South American Blastomycosis. These patients can be classified into two main clinical immunological forms.
a) mild-blastomycosis, usually of the localized forms and characterized by a low titer in complement-fixing antibody. In these forms the precipitin is the first antibody to appear in the serum of the patients and the first to disappear after treatment; b) severe-blastomycosis, clinically manifested as disseminated or generalized forms and immunologically characterized by high titer of complement-fixing antibody and precipitin reaction positive when the illness is in activity. The immunological follow up showed that the precipitin is the first antibody to disappear in the serum of these patients when they receive a good treatment. On the other hand, the precipitin is the first antibody to reappear in the circulation by the time of the occurrence of relapses of the disease. The complement-fixing antibody remains for a long time in the blood of the patients clinically cured. In the majority of cases a decrease in the titer of these antibodies can be found at the same time as the clinical improvement. The cases where the titers show a late fall, after some time of clinical improvement, are rare. The clinical relapses are accompanied by a rise in the titer of complement-fixing antibodies. The cases in which only precipitin reappears are rare and some of them have been treated again. In the evolution of a case when the titer in complement-fixing antibody is higher than 10 (by the technique of Wadsworth, Maltaner and Maltaner); or if the precipitin reaction is positive, the treatment should not be interrupted. In some cases, the titer below 5, when observed over a period of time lasting more than 6 months, may be considered a "serological scar" and the treatment can be interrupted, There are relapses that were characterized only by the serological result without clinical manifestation. Very rare cases of South American Blastomycosis do not present any precipitin or complement-fixing antibodies in the blood. In this paper, some cases have been reported with clinical and serological cure, and the true number may be higher on account of the very low number (only 10%) of localized forms registered here. The immunological study does not help so much in metastatic localized forms of South American Blastomycosis as in the nervous and adrenals localizations referred in this paper. On the other hand, cases are presented where the immunological study associated with clinical data was of much help in the diagnosis. In the study of the relatives of patients with South American Blastomycosis it was not possible to demonstrate any inter-human transmission by the precipitin and complement fixation reactions.References
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