Serodiagnosis of schistosomiasis

mesentery. A splenectomy and cholecysteromy were performed, as well as biopsies of the liver, colon, and mesenteric mass. The liver, perihilar, and colon biopsies contained multiple granulomas containing eggs of S. mansoni. There was periportal fibrosis of the liver. Microscopically, there was congestive splenomegaly and reticular hyperplasia, and a lymph node showed sinus histiocytic hyperplasia. Four months after surgery, the patient was referred to NIH for therapy. Examination revealed a firm nontender liver 10 cm below the costal margin at the midxiphoid line. The hemoglobin was 15.8 g/100 ml, hematocrit 46%, WBC 5700/mm 3 with 38% neutrophils, 46% lymphocytes, 2% monocytes, 2% basophils, and 12% eosinophils. The alkaline phosphatase was elevated at 410 U/L (range, 15-117); SGOT 76 U/L (range, 6-52); SGPT 124 IU/L (range, 2-45), total bilirubin elevated at 1.9 mg/100 ml (direct 0.3), total protein elevated at 8.5 g/100 ml, and albumin 3.8 g/100 Ixl. IgG was elevated at 322 IU/ml, IgA 248 IU/ml (range, 30--261), IgM 396 IU/ml (range, 26-266), and IgE 11,020 ng/ ml (normal value < 360 ng/mi). Chest x-ray and barium swallow was normal. The IIF test to GASP was positive, and he had an IgG response to PSAP indicating chronic infection. Stool examination revealed between 420 and 900 S. mansoni eggs/g and review of the liver biopsy revealed Symmers' fibrosis. The patient was treated with sodium antimony dimercaptosuccinate ad experienced transient arthralgia, fever, nausea, wheezing and coughing, and chest x-ray changes. At follow-up, 6 mo after treatment, no S. mansoni eggs could be detected in stools.

Comments These cases represent major disease manifestations caused by S. mansoni. As in other chronic infections, the manifestations of disease depend on a number of factors such as a history of previous exposure, intensity of exposure and infection, species or strain of schistosome, and other unknown factors. The first case is typical of acute schistosomiasis, which usually occurs in persons not previously exposed to the parasite, such as Peace Corps volunteers. This syndrome is rare in endemic populations in which initial exposure and infection goes unnoticed. Symptoms of acute schistosomiasis include fever, cough, myalgias, rash, malaise, diarrhea and weight loss. These vary but can last 2-3 mo and resolve without treatment although the patient is still infected. After years of repeated heavy exposure and infection, some persons develop Symmers' fibrosis, which is pathognomonic of severe liver involvement in schistosomiasis; the second patient demonstrates this form of the disease. There is a presinusoidal blockage resulting in portal hypertension, varices, and congestive splenomegaly. Patients commonly experience repeated bouts of hematemesis, and exsanguination is a common cause of death. Liver function is well preserved in schistosomiasis compared with alcoholic cirrhosis, as demonstrated by the second case. The liver enzymes are often slightly elevated with a normal albumin and usually without ascites. Decreased liver function does occur in schistosomiasis apparently associated with superimposed chronic active hepatitis. The second patient also demonstrates two clinical points: S. mansoni

Serodiagnosis of Schistosomiasis Shirley E. Maddison, Ph.D. Victor C. W. Tsang, Ph.D. Division of Parasitic Diseases, Center for Infectious Diseases Centers for Disease Control, Public Health Service U.S. Department of Health and Human Services Atlanta. Georgia Identification of schistosome eggs in the stool, rectal biopsy, or urine constitutes a definitive diagnosis of infec-

© 1983 by Elsevier Science Publishing Co., Inc.

tion. Detection of antibodies to schistosomes, however, is of value among U.S. expatriates upon their return to

eggs were not detected in the feces of this patient, although active granuloma were seen in tissue sections and a large number of ova were subsequently seen in the stool at NIH. Commonly, laboratories not experienced in demonstrating eggs of schistosomes fail to find eggs in stools. Also, the liver in this patient showed an enlarged left lobe, which is typical of the type of enlargement associated with S. mansoni infection. In addition to S. mansoni, there are two other species of schistosome that frequently infect man: S. haematobium and S. japonicum (as well as several other species). S. heamatobhmz causes granulomatous inflammatory lesions of the ureters and bladder. Dysuria and hematuria are common during the entire course of the infection. The signs and symptoms associated with S. japonicum resemble those of S. mansoni. For a more detailed discussion of schistosomiasis, the reader is referred to a recent review. (2). A common error in the evaluation of infected patients is to ascribe the manifestations of other diseases to incidental schistosomiasis. In most endemic populations, relatively few patients develop disease. References 1. Nash, T. E. et al. (1983). Differentiation of acute and chronic schistosomiasis by antibody responses to specific schistosome antigens. Am. J.Trop. Med. Hyg. (in press) 2. Nash, T. E. (1982). Schistosome infections in humans. Ann. Int. Med. 97:740-754. 3. Nash, T. E, E. A. Ottesen, and A. W. Cheerer. (1978). Antibody response to a polysaccharide antigen present in the schistosme gut. II. Modulation of antibody response. Am. J. Trop. Med. Hyg. 27:944-950.

the United States. In addition, positive serology may be an important lead in diagnosing patients with symptoms compatible with ectopic deposition of schistosome eggs in the skin, central nervous system, or genital tract. In addition to detection of infection, serologic assays that measure intensity of infection and the effect of chemo101

II

therapy would be advantageous to the clinician in nonendemic areas. Such assays and, in addition, an assay that could detect primary transmission or reinfection would make a valuable contribution to the evaluation of control programs in endemic areas. It is essential that assays used on an epidemiologic scale in control programs be easily performed and the relevant antigens be cost-effective and available in sufficient quantities. It is well recognized that antibody levels do not necessarily parallel intensity of infection. A more appropriate assay for indication of parasite load in schistosomiasis may prove to be the quantitation of circulating antigen. A number of investigators have detected circulating antigen or immune complexes in the serum of schistosomiasis patients. The small quantities of this antigen have necessitated radioimmunoassay, which is not well suited to seroepidemiologic studies in developing countries. With the advent of monoclonal antibodies and the development of new assays, however, the routine measurement of circulating antigen in schistosomiasis may be practical in the forseeable future. A summary of the assays and pufffled antigens discussed in this brief review are shown in Table I. The most widely used serologic assays for schis-

tosomiasis, until very recently, have involved particulate antigens. The indirect immunofluorescence assay (IIF) has been extensively evaluated (7). Frozen sections of S. mansoni adult worms are used an antigen because of interspecies crossreactivity (22). This assay, however, is quite cumbersome, subjective in reading, and quantitation in terms of serum titration is frequently not done. A modification of IIF using specific gut-associated proteoglycan antigen (GASP) in sections of adult worms prepared in Rossman's fixative (12), has been useful in differentiating acute from chronic infections. Specific antibodies of both the IgM and IgG classes to GASP are increased in acute infections (of less than 4 yr duration) compared with chronically infected groups. In the circumoval precipitin test (COPT) eggs and serum are mixed on microscope slides. Precipitates form around the eggs (6, 13). The assay is specific and sensitive, although cumbersome to perform, requires reproducibly good preparations of clean eggs, and cannot be readily quantitated. A secretory-excretory antigen consisting of polydisperse glycoproteins (PSAP) has been derived from adult S. mansoni worms (11). Use of this antigen in enzyme-linked immunosorbent assay (ELISA) or radioimmunoassay

I

T

Ill

(RIA) allows differentiation of acutely and chronically infected patients as IgM antibody response predominates in the former. Crude antigenic extracts of the various stages of the life cycle of Schistosoma spp. have resulted in assays that are below optimum both in sensitivity and specificity when used in endemic areas. The main thrust in purification of antigens has been directed to the aqueous-soluble extract of eggs (SEA) (2). However, yields of purified S. mansoni egg antigens, which include the major serologic antigen (MSA) (14), the protein antigens of Carter et al. (3), the polysaccharide antigen (PEA) (1), and the egg antigen of S. haematobium (4) and S. japonicum (8), have all been too low for extensive use. In endemic areas, antigens must be available in amounts that will permit epidemiologic surveys and must be cost effective. In preparation of SEA of eggs and adult worms of schistosomes, the amount of residual insoluble material is considerable. Therefore, we have used a systematic approach to the biochemical fractionation of schistosome proteins and glycoproteins. This included analysis of both aqueous-soluble antigens and antigens extracted from the residual particulate material with urea. Antigenic fractions from a

Table 1 Assays and Purified Soluble Antigens Assay

Antigen Used

IIF (indirect immunofluorescent assay)

(a) Frozen sections of adult worms

COPT (cirumoval precipitin test) ELISA (enzyme-linked immunosorbent assay) k-ELISA (kinetic-dependent ELISA) RIA (radioimmunoassay) EITB (enzyme-linked immunoelectro-transfer blot technique)

(b) Rossman's fixation of adult worms to demonstrate GASP Isolated eggs Soluble antigens Soluble antigens Soluble antigens Soluble antigens

Antigens GASP (gut-associated antigen from adult worms) PSAP (polydisperse glycoprotein secretory-excretory antigen, from adult worm) SEA (aqueous-soluble crude extract of eggs) MSA (major serologic antigen derived from SEA) PEA (polysaccharide antigen from eggs) MAMA (purified antigen from aqueous-insoluble, urea-extracted component of adult worms of S. mansoni)

102

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single batch of parasite material are evaluated at the same protein concentration in a kinetic-dependent ELISA (k-ELISA) (17). Serum antibody (in a standard infection serum pool) and enzyme-conjugated antiimmunoglobulin are in excess. The specific schistosome activities and crossreactivities with other parasitic infections, thus, are directly compared in any number of antigenic fractions. Sera from individuals with known infections with other parasites are used in the crossreactivity studies. S. mansoni eggs were selected for the initial systematic antigen study (21). MSA was prepared from the aqueous-soluble components according to Pelley et al. (14). The residual particulate material was extracted in 8 M urea. When compared with MSA, the urea-solubilized antigenic fraction had 20-fold greater specific schistosome activity and showed only one-sixth of the crossreactivity with trichinella. Similar results were obtained in an initial study with S. japonicum eggs (20). In the systematic fractionation of adult S. mansoni worms, the antigenic fraction (MAMA) of highest schistome activity was obtained by urea extraction of the aqueous-insoluble microsomal component (Figure 1) (18, 19). Treatment with n-butanol improved the ease of purification for this antigenic fraction. Purification on Bio Gel A50 m followed by OAE Sephadex column chromatography increased both the sensitivity and specificity. These fractionation studies have been greatly facilitated by the kELISA. Instrumentation for this assay is not yet widely available for serodiagnostic or seroepidemiologic studies. We used a conventional microplate ELISA to evaluate S. mansoni crude egg antigen SEA (2), crude adult worm antigen (9) and the purified adult worm antigen (MAMA) (18) for routine serodiagnosis. In the initial studies, excess antigen and conjugate concentrations were determined by block titrations. Unknown sera were tested in duplicate at a single dilution of 1:64. Positive and negative control sera were included in each plate. The significant serum titers in the assays with the respective anti-

© 1983 by Elsevier Science Publishing Co., Inc.

I s . mansoni: adult worms I

homogenized & Differential Centrifugation

I

|

1

Nuclear

Mitochondria

Microsome

]

Urea Treated

Urea Treated

Urea Treated

Cytosol

n-butanol treated

Bio Gel A 50m

QAE-Sephadex Figure 1. Flow chart of purification procedures that resulted in a sensitive specific microsomal antigen for adult worms of S. mansoni.

with 6% of CDC serum bank donors. The crude adult worm antigen crossreacted with sera from some amebiasis and hepatitis patients. The urea-extracted, purified MAMA did not show these crossreactivities. In an attempt to standardize the plate ELISA, a standard schistoso-

gens were derived from the mean absorbance at 490 mm plus two standard deviations of control sera from 78 healthy CDC serum bank donors. Table 2 shows that all three antigens had high levels of activity with schistosomiasis sera from endemic areas. SEA, however, showed crossreactivity

Table 2 Reactivities of S. mansoni Infection Sera and U.S. Controls in ELISA Against Crude Egg and Adult Worm, and Purified Adult Worm Antigens of S. mansoni SEA Source of Sera

Egypt Brazil Puerto Rico CDC serum bank controls Trichinosis Echinococcosis Amebiasis Hepatitis

Crude Adult

MAMA

Patients (n)

Mean OD

% Pos

Mean OD

% Pos

Mean OD

% Pos

50 49 48

1.13" 0.99 1.01

100 100 I00

1.17 0.83 1.03

100 90 96

1.17 1.15 1.09

100 98 98

78 26 21 23 65

0.24 0.52 0.33 0.22 0.26

6 50 10 0 8

0.17 0.24 0.46 0.21 0.21

0 8 67 9 8

0.11 0.25 0.20 0.09 0. ! 2

0 19 4 0 0

a Optical density of 1:64 dilution of serum. Significant level of reactivity: SEA, 0.44; crude adult, 0.39; MAMA, 0.35.

103

miasis reference serum pool was prepared in our laboratory. One microliter of the pool was arbitrarily designated as having 100 activity units. The standard serum pool is used to prepare a standard curve (10) on each ELISA plate. The amounts of the serum pool used to prepare the curve are measured in microliters. M A M A antigen and peroxidase conjugated anti-human IgG are used in excess. A typical curve is shown in Figure 2. One negative and three positive control sera are also included on each plate. Unknowns are tested in a single well of the plate by adding 200 I.tl of a 1:200 dilution of each serum (or 200 p.l of the appropriate dilution of strongly reacting sera). This allows all levels of reactivity to be determined quantitatively and results are not subject to the limitations of the ELISA reader. The activity units of the unknowns and controls are determined from the standard curve. Thus, the measurements are on a sliding scale in contrast with the two- or fourfold stepped dilutions used to determine endpoints of serum titrations. Small changes in levels of reactivity of paired sera can thus be detected. The use of a standard curve and determination of level of reactivity of sera in activity units also allows monitoring of reproducibility of the assay and the precision of the serologist. The correlation coefficient (r 2) of the standard curves in our study ranged from 0.0942 to 0.989. The coefficients of variation of repeated testing of low and high reactivity infection sera are shown in Table 3. The results using M A M A antigen and standard schistosomiasis reference serum pool are briefly summarized in Table 4. The assay shows high sensitivity and specificity. A parallel study using M A M A and standard curves from the standard schistosomiasis reference serum pool was made with the fluoroimmunoassay (FLAX) (15). In this assay, antigen is adsorbed onto cellulose acetate discs. Reaction with antibody is detected with fluorescein-conjugated antiimmunoglobulin and read in a fluorometer. The reactivities of Schistosoma infec-

I04

20

XX

1S

<

XX

XT, i .I

,

,

,

I

,

~

T

T

I

5,

,

,

10

~

,

[ 1S

ABSORBANCE 490ram

Figure 2. Standard curve prepared in duplicate with standard serum pool in each ELISA plate sensitized with MAMA antigen. Serum pool added in microliter amounts. (X = read-

ings for each amount of serum pool; • = calculated best fit curve.)

tion sera were remarkably comparable to those obtained in ELISA, but low levels of crossreactivity were observed with the ascariasis/trichiuriasis sera and with the paragonimiasis and malaria sera. These were all negative in ELISA. The cause of this low-level crossreactivity in FlAX remains unclear. It does, however, preclude the use of FlAX for routine serodiagnosis

under the conditions used in our laboratory. Monoclonal antibodies have been used in our laboratory for immunoaffinity purification of antigens from the different life cycle stages of S. mansoni. The reactivity of these monoclonal antibodies can be well defined by studying them in the recently introduced enzyme-linked immunoelectro-

Table 3 Run-to-Run Variation of Low- and High-Reactivity Schistosomiasis Sera in ELISA Using MAMA Antigen and a Standard Curve Ser,Lm

NT

X

SD

% CV

Low

22

l0

1.0

10.2

High

24

84.5

10.4

12.2

NT, not tested; X, mean activity units per ltl; % CV, percent coefficient of variation.

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Table 4 Reactivities of S. m a n s o n i Infection Sera and U.S. Controls Against Purified Adult W o r m Antigen (MAMA)

in ELISA Employing a Standard Curve Source of Sera

Patients (n)

Egypt Brazil Venezuela Puerto Rico U.S. Expatriates CDC serum bank controls Trichinosis Ascariasis and/or trichiuriasis Echinococcosis Amebiasis Hepatitis

50 43 34 46 11

Mean Activity Units

171" 34 58 76 64

% Pos

6-1816 1-146 3--277 10--322 19-258

94 74 94 100 100

81 24

0.4 0.9

0.2--0.8 0-2.1

0 0

13 24 24 62

5.0 1.6 0.1 0.4

1.4-7.2 0-6.8 0-0.5 0.2-0.9

0 0 0 0

a Activity units per ptl. transfer blot technique (EITB) (16). This assay enables the visualization, following sodium dodecyl sulphatepolyacrylamide electrophoresis, of those particular antigenic bands that react with sera or monoclonal antibodies. Similarly, an affinity purified antigen can be compared with its parent antigen. Because loss of circumoval precipitin reactivity is well recognized as indicating cure following chemotherapy for schistosomiasis, we used a COPTreactive monoclonal antibody to isolate an antigen from the urea-extracted fraction of S. mansoni eggs. The ac-

Significant level of reactivity: ~>10 activity units per p.l. tivity in the COPT antigen appears to be mainly in the high molecular weight components and is probably glycoprotein. Possibly, multiple epitopes participate in the COPT; Hillyer (5) recently showed that MSA (2) had epitope(s) that induced a COPT-reactive antibody in immunized mice. Preliminary studies with our soluble COPT antigen of infection sera, before and after treatment, indicate it measures antibodies different from those detected by MAMA. In addition, reactivity to the COPT antigen in plate ELISA following treatment decreased considerably earlier than did reactivity

References 1. Boctor, F. N., T. E. Nash and

A. W. Cheever. (1979). Isolation of a polysaccharide antigen from Schistosoma mansoni eggs. Am. J. Trop. Med. Hyg. 122:39-43. 2. Boros, D. and K. S. Warren. (1970). Delayed hypersensitivity-type granuloma formation and dermal reaction induced and elicited by a soluble factor isolated from Schistosoma mansoni eggs. J. Exp. Med. 132:488-507. 3. Carter, C. E. and D. G. Colley. (1979). Partial purification and characterization of Schistosoma mansoni soluble egg antigen with Con A-sepharose Chromatography. J. Immunol. 122:2204-2209. 4. Hamburger, J. et al. (1982). Analysis and preliminary purification of glycoproteins isolated from eggs in the urine of patients with Schistosoma

© I983 by Elsevier Scieace Publ~shlngCo., Inc.

Range

to MAMA. Whether this antigen will be useful in assessing the effect of chemotherapy on an epidemiologic scale remains to be evaluated. Application of new antigen preparation procedures and sensitive quantitative assays by a number of investigators are resulting in rapid progress towards sensitive, species-specific serodiagnostie capability. Seroepidemiology will contribute to the evaluation of schistosomiasis control programs when serologic assays that can detect recent transmission and the efficacy of chemotherapy have been developed.

haematobium infection. J. Immunol.

5.

6.

7.

8.

129:1711-1714. Hillyer G. V. and R. R. Pelley. (1980). The major serological antigen (MSA) from Schistosoma mansoni eggs is a "circumoval" precipitinogen. Am. J. Trop. Med. Hyg. 29:582-583. Hillyer, G. V. et ai. (1979). Immunodiagnosis of infections with Schistosoma mansoni: Comparison of ELISA, radioimmunoassay, and precipitation tests performed with antigens from eggs. Am. J. Trop. Med. Hyg. 28:661-669. Kagan, I. G. (1980). Serodiagnosis of parasitic diseases. In E. H. Lennett et al. (eds), Manual of Clinical Microbiology. American Society of Microbiologists, Washington, D.C. Long, G. W. et al. (1982). Efficacy of purified Schistosoma japonicum egg antigens for ELISA serodiagnosis of

9.

10.

11.

12.

human schistosomias japonica: Specificity and sensitivity. Am. J. Trop. Med. Hyg. 31:1006--1014. Lunde, M. N., E. A. Ottesen, and A, W. Cheerer. (1979). Serologic differences between acute and chronic schistosomiasis mansoni detected by enzyme-linked immunosorbent assay (ELISA). Am. J. Trop. Med. Hyg. 28:87-91. Mondesire, R. R., D. E. Charlton, and I. R. Tizard. (1981). A standardized enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to Toxoplasma gondii. J. Immunoassay 2:45-57. Nash, T. E. et al. (1983). Differentiation of acute and chronic schistosomiasis by antibody responses to specific schistosome antigen. Am. J. Trop. Med. Hyg. (in press) Nash, T. E., E. A. Ottesen and

105

13.

14.

15.

16.

A. W. Cheever. (1978). Antibody response to polysaccharide antigen present in the schistosome gut II modulation of antibody response. Am. J. Trop. Med. Hyg. 27:944-950. Oliver-Gonzales, J. (1956). Anti-egg precipitins in the serum of humans infected with Sehistosoma mansoni. J. Infect. Dis. 95:86-91. Pelley, R. P. et al. (1976). Schistosoma mansoni antigens I. Identification and purification of three major antigens and the employment of radioimmunoassay for their further characterization. J. Immunol. 117:1553-1560. Slemenda, S. B., M. liitchings, and S. E. Maddison. (1980). Standardization of FIAX~ for schistosomiasis using crude cercarial and adult worm antigens. J. Parasitol. 66:893-897. Tsang, V. C. W., J. M. Peralta, and A. R. Simons. (1983). Enzyme-

Some Recent Experiences in Serodiagnosis of Schistosomiasis in the United States Shirley E. Maddison, Ph.D., Victor C. W. Tsang, Ph.D. Robert E. Fontaine, M.D. Division of Parasitic Diseases, Center for Infectious Diseases Centers for Disease Control. Public Health Service u . S . Department o f Health and Human Sen'ices Atlanta, Georgia

G. R. Istre, M.D. Division of Field Services, Epidemiology Program Office EIS Officer at Oklahoma Department of Health Oklahoma City, Oklahoma

In the continental United States, the indirect immunofluorescence assay (IIF) (8), using frozen sections of adult worms of Schistosoma mansoni, is the most extensively evaluated (1) test for routine detection of antibodies to schistosomes. The circumoval precipitin test (5), and an IIF, designed to show reaction with carbohydrate-rich material in the gut of adult worms (4) are also used. In addition, a number of modifications o f the quantitative enzyme-linked immunoabsorbent assay (ELISA) (7) are being evaluated. It is

106

linked immunoelectrotransfer blot techniques (EITB) for studying the specificities of antigens and antibodies separated by gel electrophoresis. Meth. Enzymol. 92:377-391. 17. Tsang, V. C. W., B. C. Wilson, and J. M. Peralta, (1983). Quantitative, single-tube, kinetic-dependent enzymelinked immunosorbent assay (kELISA). Meth. Enzymol. 92:391-403. 18. Tsang, V. C. W. et al. (1983). Schistosoma mansoni adult microsomal antigens, a serologic reagent 1. Systematic fractionation, quantitation and characterization of antigenic components. J. Immunol. 130:1359-1365. 19. Tsang, V. C. W. et al. (1983). Schistosoma mansoni adult microsomal antigens, a serologic reagent. II. Specificity of antibody responses to the S. mansoni microsomal antigen (MAMA). J. Immunol. 130:1366-1370.

20. Tsang, V. C. W. et ai. (1982). Fractionation and quantitation of egg antigens from Schistosoma japonicuma by the single-tube kinetic-dependent, enzyme-linked immunosorbent assay (kELISA): Higher antigenic activity in urea-soluble than in aqueous-soluble fractions. J. Parasitol. 68:1034-1043. 21. Tsang, V. C. W., X. Tao, and S. E. Maddison. (1981). Systematic fractionation of Schistosoma mansoni ureasoluble egg antigens and their evaluations by the single-tube, kinetic-dependent, enzyme-linked immunosorbent assay (k-ELISA). J. Parasitol. 67: 340-350. 22. Wilson, M., A. J. Sulzer, and K. W. Walls. (1976). Modified antigens in the indirect immunofluorescence test for schistosomiasis. Am. J. Trop. Med. Hyg. 23:1072-1076.

well recognized that the antigens of the various stages (adult worm, egg, and cercaria) have multiple common antigens; similarly, crude extracts of each stage of the various species of Schistosoma crossreact. Crossreactivity, therefore, is evident in IIF. Thus, speciation has not been possible in routine serodiagnosis. Differentiation of S. mansoni and S. haematobium infections would be of value as S. haematobium is not affected by the drugs commonly used for treatment o f S. mansoni infections. Schistosomiasis was diagnosed in 5 of 10 U.S. tourists who had been exposed to Schistosoma-contaminated river water in Africa. In retrospect, the index case presented with Katayama syndrome (the hallmark of early schistosomiasis) with an onset of approximately 3 wk following infection. Diagnosis of S. mansoni infection was not made until the patient was reevaluated, approximately 1 mo later. At this time, S. mansoni eggs were identified in the feces o f the patient, and serology for schistosomiasis was positive by IIF. As a result of these findings, all other persons who had similar exposure to contaminated water were contacted. Four had developed generally mild clinical symptoms and signs compatible with acute schistosomiasis. All persons with symptoms had low fecal

egg counts and positive serology. Only one of five asymptomatic persons had eggs in the stool and a detectable antibody response. The IIF was postitive in the five infected individuals (Table I). Although the egg counts were low, the quantitative plate ELISA (3) with a purified microsomal antigen from adult worms of S. mansoni (MAMA) (2) showed

Table I Results of Serologic Assays on 11 Participants in a 1981 O m o River Rafting Expedition

Patient

lip

Plate ELISA b with MAMA c Antigen

1 2 3 4 5 6

+ + + + + +

26a 131 27 40 45. 19

5 uninfected

-

0.31--0.65

° IIF, indirect immunofluorescence with frozen sections of adult worms of S. mansoni. b ELISA, enzyme-linked immunoadsorption assay. c MAMA, purified microsomal antigen from adult worms of S. mansoni. a Activity units per ml; significant level = 10 activity units.

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