Clinical Comparison of Two Mexican Coccidioidins

Coccidioidin, an extract from the saprophytic mycelial form of Coccidioides spp., has been a very useful antigen preparation both for skin and serological tests for coccidioidomycosis. Unfortunately, coccidioidin is not currently available for skin testing in the United States. Coccidioidin has been produced commercially in Mexico by a vaccine and reagents laboratory of the Mexican Federal Government. It also has been produced at the Microbiology Laboratory of the Faculty of Medicine, Universidad Nacional Autónoma de México exclusively as an antigen for research projects. The objective of the study was to compare both coccidioidins in their reactivity and safety when applied in humans. One hundred and eighty-four volunteers were tested; median age was 33 (range 14–82). When the cutoff point is set in 5 mm, 88 subjects (47.8%) had a positive test for the commercial coccidioidin and 76 (41.3%; CI_95% 0.50, 1.15; P = 0.20) were positive with the research antigen. Seventy-five subjects were positive for both antigens and 96 were negative for both. Fifty-nine subjects (31.3%) reported an adverse reaction after the application of the antigen; they were mostly very mild local reactions. Mexican research coccidioidin is a safe and reliable antigen that can be used for the detection of coccidioidomycosis infection in mammals.     

THE DISTRIBUTION OF COCCIDIOIDOMYCOSIS IN SOUTHERN CALIFORNIA

In coccidioidin skin test surveys among persons of high school age in Saugus, Canoga Park, Banning and Palm Springs areas the average incidence of positive reaction was 15 per cent. Although considerably less than the 68 per cent incidence reported among high school students of Kern County, it is high enough to indicate pockets of relatively high endemicity in Southern California below the San Joaquin Valley.Histoplasmin tests were performed on most of the persons tested with coccidioidin in this survey. The over-all incidence of positive reaction in the group was 7.6 per cent. Most of the subjects with positive reaction to histoplasmin gave a history of having previously lived in some area in the central United States where histoplasmosis is known to be endemic.A few subjects who had positive reaction to coccidioidin tests and who had lived in areas known to be endemic for coccidioidomycosis but not for histoplasmosis, also had positive reaction to histoplasmin. However, the induration produced was always smaller than that caused by the coccidioidin reaction, and there was minimal confusion in interpreting the tests.     

A Reformulated Spherule-Derived Coccidioidin (Spherusol) to Detect Delayed-Type Hypersensitivity in Coccidioidomycosis

The ability of spherule-derived coccidioidin containing 0.4?% phenol and 0.0001?% thimerosal in buffered saline to induce delayed-type hypersensitivity (DTH) was evaluated in four separate studies. The skin test antigen was titrated in 20 adult volunteers with a recent history of pulmonary coccidioidomycosis using intradermal doses of 0.4, 0.8, and 1.6?μg of antigen, based on total dry weight. Based on these data, a dose of 1.27?μg was shown to elicit a mean?±?SEM induration response of 23.5?±?2.3?mm at 48?h, similar to the 23.6-mm response after 48?h of the U. S. Reference coccidioidin last tested approximately 13?years ago. The 1.27?μg dose in 0.1?mL of the spherule-derived antigen (Spherusol) was then examined in three separate groups of adult volunteers to determine the sensitivity and specificity of the product. Fifty-nine of 60 individuals living in a non-endemic area for coccidioidomycosis were skin test negative to Spherusol. Twelve subjects with a recent history of pulmonary histoplasmosis were skin test negative to Spherusol. Finally, 51 of 52 individuals with a recent diagnosis of acute pulmonary coccidioidomycosis were skin test positive to Spherusol. Within this group, prior therapy with fluconazole did not appear to reduce the reactivity to Spherusol. No serious adverse events were observed in the four studies. From these data, Spherusol was found to be safe and has an overall observed sensitivity and specificity of ≥98?% in detecting DTH in coccidioidomycosis.     

Intradermal reactions with coccidioidins in different towns of San Luis Province

Intradermal tests were used to determine the extent of the endemic zone of coccidioidomycosis in Argentina. We performed our endemiological study among school-aged children and grown-ups in San Luis city and in the following towns: Nogoli, Villa de la Quebrada, Balde, Salinas del Bebedero, Beazley, Fraga and Eleodoro Lobos. We employed three coccidioidins for each person--Negroni's coccidioidin, coloured coccidioidin and uncoloured coccidioidin. Reactions were usually read 48 h after inoculation. After inoculating 1,262 individuals we could read only 1,069 results. Overall, the positive reactors to one, two or three coccidioidins were 14.8%. We found no relation between positive reactors and the sex of subjects, but every group showed a noticeable relation between age and positive reactions, especially in older people. The positive reactors for each of the coccidioidins were as follows: Negroni coccidioidin: 10.1%: coloured coccidioidin: 4.9% and uncoloured coccidioidin: 5.0%.     

DIAGNOSIS OF PULMONARY COCCIDIOIDAL INFECTIONS

A wide variety of pulmonary lesions may be caused by coccidioidomycosis. Suspicion of coccidioidomycosis may be substantiated by careful clinical-epidemiological histories. The first laboratory procedure should be a coccidioidin skin test. If the reaction to the test is positive, serological tests are next. Also, if there is no reaction to coccidioidin, serological tests are still indicated if dissemination is suspected. The more severe the infection, the greater the probability of establishing a diagnosis serologically. In only three-fifths of patients with coccidioidal cavities can the diagnosis be fixed serologically. In such patients if differential skin tests are not conclusive, attempt should be made to recover the fungus. However, this is accompanied by great risk of laboratory infection. Eosinophilia and accelerated erythrocyte sedimentation are only circumstantial items of evidence, as is the appearance of the pulmonary roentgenogram.     

An Archived Lot of Coccidioidin Induces Specific Coccidioidal Delayed-type Hypersensitivity and Correlates with in vitro Assays of Coccidioidal Cellular Immune Response

No test for assessing cellular immune response in coccidioidomycosis is currently available in the United States. In the present study, we tested 49 healthy subjects living in the coccidioidal endemic region with a 1:55.8 dilution of a single lot of coccidioidin archived since the 1970s. In this group, 23 evaluable subjects demonstrated ≥5 mm of induration at 24, 48 or 72 h, with a mean±SEM maximum induration of 18.4±4.0 mm. The induration results among 14 subjects reactive at 24 h were compared to those from 179 individuals in an historical cohort studied in the 1980s using a reference lot of coccidioidin. Results were within 5% and not significantly different (P=0.924). The maximum induration response of all evaluable subjects correlated significantly with the results of in vitro tests of coccidioidal cellular immunity using supernatant interferon-gamma concentration and CD69 expression on T cells (Spearman rank correlation coefficients 0.69 and 0.68, respectively; P<0.01 for both). These data suggest that archived coccidioidin retains its potency and specificity and that in vitro test of coccidioidal immunity may have utility in the measurement of coccidioidal cellular immunity.     

Immunology of histoplasmosis: Humoral and cellular activity from a polysacchari-de-protein complex and its deproteinized fraction in experimentally immunized mice

In a previous publication it was reported that a polysaccharide-protein complex (PPC), sensitive to -glucosidase, was isolated from Histoplasma capsulatum. This complex was strongly reactive in an agar gel diffusion assay with sera from patients with histoplasmosis, but was unreactive with sera from patients with coccidioidomycosis. Here, the studies with human sera have been expanded and attempts were made to determine the response of mice immunized with nonviable H. capsulatum or Cocccidioides immitis to PPC or its deproteinized fraction (D-PPC) using more sensitive tests for antibody and including also test for cell-mediated immunity. Histoplasmin and coccidioidin were compared with PPC or its deproteinized fraction (D-PPC) in all assays. In a counterimmunoelectrophoresis (CIE) assay, PPC and D-PPC reacted only with sera from patients with histoplasmosis, whereas cross reactions were noted with histoplasmin and coccidioidin using heterologous sera. Cross-reaction were observed with all four antigen preparations and both types of antisera using a micro complement fixation assay. The assay for macrophage migration inhibitory factor (MIF) was also relatively nonspecific, in that inhibition occurred with cells from animals sensitized with Histoplasma or Coccidioides using both homologous and heterologous antigens. In the footpad assay, histoplasmin and coccidioidin were highly cross-reactive in animals sensitized with the heterologous fungus, but the PPC and D-PPC from H. capsulatum elicited significant reactions only in animals sensitized with Histoplasma.     

Value of the Concomitant Use of Complement Fixation and Immunodiffusion Tests in the Diagnosis of Coccidioidomycosis

Complement fixation titers of 1:2 and 1:4 with coccidioidin are not always of diagnostic significance. The concurrent use of the complement fixation and immunodiffusion tests is an effective means for specific serologic diagnosis of coccidioidomycosis in patients with low levels of complement-fixing antibodies. In routine testing, it is recommended that only sera that are negative for complement-fixing antibodies at 1:8 but positive in the immunodiffusion test with coccidioidin be selected for titration at the 1:2 and 1:4 levels.     

Results of skin test surveys for systemic Mycoses in Iran

Several investigators had carried out histoplasmin skin test surveys in 3 different areas of Iran before 1960 (5). To detect skin sensitivity to both histoplasmin and coccidioidin a study was carried out by the Institute of Public Health Research in parts of 7 of the country's 13 provinces on both apparently healthy persons and on clinic patients from February 1962–September 1966. These studies uncovered a small focus with a rather high level of positive reaction to histoplasmin (about 17 %). Positive cultures ofHistoplasma capsulatum were not obtained from either sputum of the positive reactors or from collected soils. In spite of a previously reported case of coccidioidomycosis, very low percentages of positive reaction to coccidioidin were noted.These studies were supported in part by the Institute of Public Health Research Teheran University, and funds of the Ministry of Health and Plan Organization for Project No. 631101 and N.I.H. Grant No. TW00170.Paper read at the Eighth International Congresses for Tropical Medicine and Malaria, September 1968, Teheran (Iran).     

Spectrum of in vivo and in vitro cell-mediated immune responses in coccidioidomycosis.

The cell-mediated immune responses of 12 healthy, coccidioidin skin-test positive subjects (Group I) were compared with those of 15 healthy, coccidioidin skin-test positive persons who had primary asymptomatic coccidiodomycosis, (Group II), 12 patients with active, pulmonary coccidioidomycosis (Group III), four patients with disseminated disease (Group IV), and five patients who had been in clinical remission for 1 year or longer (Group V). Lymphocytes from healthy subjects in Groups I and II responded in vitro to Coccidioides immitis antigen by undergoing an increased DNA synthesis (lymphocyte transformation) and/or by producing macrophage migration inhibitory factor (MIF). In contrast, patients in Groups III and IV failed to respond to Coccidioides antigens in vivo (skin tests) or in vitro (lymphocyte transformation and production of MIF). The responses of subjects in Group V with inactive disease fell in between those of healthy donors in Groups I and II and patients in Groups III and IV. The cellular immune defect, in terms of antigen recognition, appeared to be specific for C. immitis in all but one patient.     

Coccidioidomycosis: Evaluation of parameters used to predict outcome with amphotericin B therapy

A literature review of case histories describing the use of amphotericin B for the treatment of disseminated coccidioidomycosis was performed to detect parameters that were predictive of therapeutic outcome. Several factors were significantly different between patients who were well during prolonged follow-up versus those with active or recurrent disease: 1) mean complement fixation (CF) titer before treatment was lower in patients who were well; 2) well patients had a greater magnitude fall in CF titer during amphotericin B therapy; 3) mean CF titer after amphotericin B treatment was lower in patients who were well; and 4) patients with a positive coccidioidin skin test before therapy were more likely to be well at 6 months. There was no correlation between total amphotericin B dose or duration of therapy and therapeutic outcome.     

Coccidioidomycosis in San Fernando Valley; report of a study carried out in 1951-1952

Four cases of benign pulmonary coccidioidomycosis occurred during May 1951 among some 35 teenage students at a boarding school and farm for boys that occupies 30 acres in the northwestern section of the San Fernando Valley within the City of Los Angeles. Epidemiological and serological study of the patients yielded evidence that exposure had occurred on the farm or nearby. Correlation of results of skin testing for sensitivity to coccidioidin among students over the subsequent months and of serologic studies in cases in which there was positive reaction to skin tests indicated exposure to coccidioides during the time the subjects were in residence at the school. In addition, two boys had conversion from negative to positive reaction to coccidioidin while they were in residence.     

COCCIDIOIDOMYCOSIS IN SAN FERNANDO VALLEY—Report of a Study Carried Out in 1951-1952

Four cases of benign pulmonary coccidioidomycosis occurred during May 1951 among some 35 teenage students at a boarding school and farm for boys that occupies 30 acres in the northwestern section of the San Fernando Valley within the City of Los Angeles.Epidemiological and serological study of the patients yielded evidence that exposure had occurred on the farm or nearby.Correlation of results of skin testing for sensitivity to coccidioidin among students over the subsequent months and of serologic studies in cases in which there was positive reaction to skin tests indicated exposure to coccidioides during the time the subjects were in residence at the school. In addition, two boys had conversion from negative to positive reaction to coccidioidin while they were in residence.     

Coccidioidomycosis: Early Immunologic Findings

T and B lymphocyte number and lymphocyte response to phytohemagglutinin, pokeweed mitogen, concanavalin A, coccidioidin and streptokinase-streptodornase (SKSD), plus monocyte ingestion of coccidioidin- and IgG-coated chicken erythrocytes were measured in 5 patients with coccidioidal meningitis, 11 with nonmeningeal extrapulmonary cocidioidomycosis and 5 with localized pulmonary infections. These cases were evaluated within six months of the onset of infection. Lymphocytic responses to phytohemagglutinin, pokeweed mitogen, concanavalin A, SKSD and coccidioidin and monocytic ingestion of coccidioidin- and IgG-coated chicken erythrocytes were severely decreased in patients with meningeal and nonmeningeal, extrapulmonary coccidioidomycosis but not in patients with localized pulmonary infections. T and B cell numbers, however, were normal in all groups. Thus, defects in cellular immunity are involved in the pathogenesis of extrapulmonary coccidioidomycosis and measurements of lymphocytic and monocytic function may identify patients prone to extrapulmonary infection.     

Positive Fluorescent Treponemal Antibody Reactions in Diabetes

In a survey of 129 diabetic patients and 142 normal individuals, a significantly higher percentage of positive reactions in the fluorescent treponemal antibody-200 (FTA-200) test was found among diabetic patients than in the normal population. Absorption of all FTA-200-reactive sera with an extract of Reiter's treponeme eliminated most of the positive reactions in sera from diabetic patients, and three of the five positive reactions detected in sera from apparently normal subjects. On immunoelectrophoresis, precipitin bands developed most frequently between the Reiter sorbent and sera from diabetic patients positive in the FTA-200 test. Serum components responsible for FTA reactivity and precipitin reactions against the sorbent were resistant to treatment with mercaptoethanol, suggesting antibody of the IgG class. Cross-reacting antibodies produced in response to normal treponemal flora, and perhaps acquiring enhanced reactivity by means of nonspecific interacting substances in sera peculiar to the altered physiological state of diabetes, are suggested as possible causes of positive reactions of unabsorbed sera. No correlation could be made between age of the diabetic patient, treatment or duration of the disease, and FTA or precipitin reactivity of the patient's serum.     

Measurement of Cellular Immunity in Human Coccidioidomycosis

Measurement of cellular immune response in human coccidioidomycosis, a disease of the Western Hemisphere caused by the soil-dwelling fungus Coccidioides, began more than 75 years ago. Numerous studies have shown that measurement of coccidioidal cellular immunity is a useful epidemiologic and clinical tool. The first widely used coccidioidal skin-test reagent, coccidioidin, was derived from a filtrate of cultured mycelia of Coccidioides by Smith and colleagues in the 1940's. It remained the standard until the 1970's, when spherulin, obtained from the spherule form of the fungus, was found to be more sensitive. Both reagents are specific indicators of coccidioidal cellular immunity. Since then, other coccidioidal antigens have been identified and in vitro tests of cellular immunity have been shown to yield comparable results to skin testing. In vitro assays have also begun to open a window on the specific immunologic events of coccidioidal cellular immunity. Persistent expression of coccidioidal cellular immune response appears to augur an improved clinical outcome. Despite this, a study of a coccidioidal vaccine failed to demonstrate efficacy. Current and future studies are focused on modulating the coccidioidal immune response in vitro and in vivo and on developing an improved vaccine.     

Immunologic and clinical improvement of progressive coccidioidomycosis following administration of transfer factor

Three patients with progressive coccidioidomycosis were given preparations of transfer factor (TF). Adverse reactions to TF were minimal. Following TF administration two of these patients had prolonged clinical remissions in their coccidioidal disease. Cellular immune responses were sequentially evaluated by coccidioidininduced delayed-type skin tests, lymphocyte blast transformation and macrophage inhibition factor production (MIF). These three patients each exhibited different cellular immune patterns before and after TF administration. Two patients converted their coccidioidin skin tests, and one converted lymphocyte transformation response to coccidioidin. Also, TF apparently favorably affected the MIF response in all three patients.     

Coccidioidomycosis in South America. A review of its epidemiology and geographic distribution

Argentina. Twenty-seven human cases and coccidioidin skin-test surveys have located the endemic area of coccidioidomycosis between the 27th and 40th south parallels. Climate is of the arid steppe type in the southern zone, arid hill and prairie in the intermediate zone, and arid hill and prairie plus hot tropical in the northern zone. Temperature ranges from 5° C to 29° C, vegetation is xerophytic and annual rainfall is from 300 to 500 mm. Paraguay. On the basis of human cases and coccidioidin surveys, the endemic area has been delimited between the 19th and 24th south parallels. It was a hot, dry, windy climate with temperature reaching 45° C, an annual rainfall average of 500 mm and xerophytic vegetation. Colombia. On the basis of two human cases and coccidioidin test surveys, an endemic area of low prevalence was confirmed in the northeast between the 10th and 12th north parallels. Altitude in this region is from 2 to 300 meters above sea level, temperature averages about 29° C. Within this region two different areas can be differentiated — one in the north where vegetation is tropical desert brush type and rainfall ranges between 125 and 500 mm; the second in the south with grass and cotton culture and rainfall from 500 to 2000 mm. Venezuela. Thirty-five human cases and nearly 60,000 skin tests made from east to west in the northern part of the country, where the population is concentrated, showed that the endemic area is situated between the 9th and 12th north parallels. This is an arid region with desert soils. Altitude ranges from sea level to 800 meters, annual temperature averages 24° C and rainfall 500 mm in some places, and 29° C and less than 400 mm in others. More than 172 species of plants have been identified in the zone but cacti predominate.C. immitis was isolated from soil collected at a site where a patient had become infected. Bolivia, Peru and Ecuador.Mackinnon studied a patient coming from Bolivia, but he has expressed doubt about the Bolivian origin of the infection because the patient had lived in the Paraguayan Chaco the previous year. More information is necessary to evaluate the human case mentioned in Peru byBinder. Cases reported from Ecuador appear to have been paracoccidioidomycosis and leishmaniasis rather than coccidioidomycosis.Many species of rodents and other wild and domestic animals share with man the possibility of infection in the four countries where the endemic areas have been confirmed.Paper read at the Eighth International Congresses for Tropical Medicine and Malaria, September 1968, Teheran (Iran).     

Agar-Gel Precipitin-Inhibition Test for Coccidioidomycosis: I. Preliminary Evaluation of the Complement-Fixation and Agar-Gel Precipitin Tests in the Serodiagnosis of Human Coccidioidomycosis

The agar-gel precipitin-inhibition serological test for coccidioidomycosis was a more sensitive indicator of Coccidioides immitis antibodies than the tube precipitin, the agar-gel immunodiffusion, in the complement-fixation tests in assaying monkey sera, whether these sera were from prechallenge-vaccinated or postchallenged animals. When applying this technique to the assay of human sera, an analogous finding generally persisted. However, some human sera were positive by the complement-fixation test and negative by the agar-gel precipitin-inhibition test. These sera were diffused in agar-gel against various coccidioidin complement-fixation, tube precipitin, and agar-gel precipitin-inhibition test antigens with essentially negative results.     

Serologic Detection of Coccidioidomycosis Antibodies in Northeast Brazil

Coccidioidomycosis is a systemic infection caused by Coccidioides spp. The disease is endemic in Brazil but its incidence is underreported as it is not a notifiable disease. This article presents the results of a serologic survey carried out with 229 volunteers in northeast Brazil by the immunodiffusion (ID) test with commercial Coccidioides spp. antigens. The commercial ID test detected 15 individuals without clinical diagnosis of the disease and two individuals in treatment for coccidioidomycosis. Regarding the epidemiological data, most of the positive individuals were male, between 18 and 65 years of age and were engaged in armadillo hunting. Three women who had never participated in armadillo hunts also displayed positive results for coccidioidal antibodies. Besides armadillo hunts, exposure to environmental dust in endemic areas may account for the serologic response detected in the study. The data from this study suggest the importance of performing epidemiological surveys for coccidioidomycosis in order to understand the prevalence of this disease in Brazil.