Fluctuation of schistosome circulating antigen levels in urine of individuals with Schistosoma mansoni infection in Burundi.

We studied the fluctuations of schistosome circulating antigens in urine as compared with fecal egg counts in 60 Burundese individuals infected with Schistosoma mansoni. Levels of circulating anodic antigen (CAA) and circulating cathodic antigen (CCA) in the urine were determined by quantitative enzyme-linked immunosorbent assays. Fecal samples were simultaneously collected and examined with duplicate Kato-Katz slides. Significant correlations were consistently found between circulating antigen levels in urine and fecal egg counts. Although both antigen levels and egg output fluctuated, there was less fluctuation of CCA levels in urine than of fecal egg counts. All individuals had CCA in at least one urine sample and 82% were at least once positive for egg counts. Positive CCA levels were found in at least one urine sample in 75% of all individuals, but levels were low. Our results show that detection of CCA in urine is a sensitive, quantitative, and reliable method for noninvasive diagnosis and screening of S. mansoni infections, due to the relatively low fluctuations.

Day-to-day variation of egg output and schistosome circulating antigens in urine of Schistosoma haematobium-infected school children from Gabon and follow-up after chemotherapy.

A group of 31 school children from Gabon infected with Schistosoma haematobium was examined before praziquantel therapy and followed on days 3, 9, 14, 21, 24, 28, 31 and 35 after therapy. The day-to-day variation of schistosome circulating antigens, urinary egg output, and the reagent strip index (RSI, a pathologic marker) was studied in six consecutive pretreatment urine samples collected under standardized conditions. The geometric mean pretreatment for egg output ranged between 97 and 223 eggs per 10 ml of urine; for urine circulating anodic antigen (CAA) levels this was 44-154 pg/ml and for circulating cathodic antigen (CCA) levels this was 180-601 pg/ml. On the day of treatment, 97% of the children had viable eggs in their urine, 87% had a positive RSI; positive CAA levels were detected in 61% of the children and positive CCA levels in 77%. A significant correlation was consistently found between RSI and egg counts. Five weeks after chemotherapy egg output, levels of CAA and CCA and the severity of pathologic findings in the lower renal tract, as indicated by the RSI, had decreased significantly in all cases. Our results indicate that egg output in urine is an accurate method for diagnosis of S. haematobium, and additionally show less day-to-day variation than detection by ELISA of schistosome circulating antigens in urine.

Presence of the schistosome circulating anodic antigen (CAA) in urine of patients with Schistosoma mansoni or S. haematobium infections.

We investigated the presence of the circulating anodic antigen (CAA) in the urine of schistosomiasis patients. This genus specific antigen was hitherto demonstrated only in the serum of schistosomiasis patients. The urine of 80 patients with Schistosoma mansoni infections, 33 patients with S. haematobium infections, and 2 patients with mixed S. haematobium and S. mansoni infections were screened by a quantitative enzyme-linked immunosorbent assay (ELISA). CAA was demonstrated in 81% of those with intestinal schistosomiasis and in 97% of those with urinary schistosomiasis. CAA titers were less than 1:0.2-1:51.2. Results were compared with circulating cathodic antigen (CCA) titers in urine obtained in an indirect hemagglutination assay (IHA). CCA was generally not detectable in the urine of patients with S. haematobium infection, but was demonstrated in the urine of 85% of the patients with S. mansoni infection. Both CAA titers and CCA titers correlated positively with the number of S. mansoni eggs excreted in the feces, but CAA titers did not show a significant correlation with the number of S. haematobium eggs in urine. Both antigen titers showed a moderate correlation with the serum CAA level in schistosomiasis mansoni. The discovery of CAA in the urine of the majority of schistosomiasis patients tested suggests the use of urine samples for non-invasive immunodiagnosis of the disease.

Improved diagnostic performance of the circulating antigen assay in human schistosomiasis by parallel testing for circulating anodic and cathodic antigens in serum and urine.

Serum and urine levels of two Schistosoma circulating antigens, the circulating anodic antigen (CAA) and the circulating cathodic antigen (CCA), were determined by monoclonal antibody-based enzyme-linked immunosorbent assays in 56 Egyptian patients infected with S. mansoni and in 12 patients infected with both S. mansoni and S. haematobium. Both CAA and CCA could be specifically demonstrated in 82% and 88% of the serum samples and in 88% and 87% of the urine samples, respectively. While complete specificity was maintained, sensitivity was increased to a range of 91-98% by parallel use of the two circulating antigen assays, i.e., an individual with a positive titer for at least one of the assays was considered to be infected. A combination of CAA and CCA determinations in urine samples only resulted in a sensitivity of 94%. However, the highest sensitivity was achieved when the serum-CCA assay was combined with the urine-CCA assay (98%) or with the urine-CAA assay (97%). Sensitivity could not be increased further by combining more than two tests. A significant correlation was demonstrated between the level of circulating antigen and the number of parasite eggs in feces in each of the four assays. In addition, the levels of CAA and CCA in serum and urine were significantly correlated with each other. Our results indicate that diagnosis of schistosome infections by detection of circulating antigens can be significantly improved by parallel testing for multiple antigens.

Quantitative determination of circulating soluble egg antigen in urine and serum of Schistosoma mansoni-infected individuals using a combined two-site enzyme-linked immunosorbent assay.

Two enzyme-linked immunosorbent assays (ELISA-5B1 using monoclonal antibody [MAb] 114-5B1-A [IgG1] and ELISA-4D12 using MAb 114-4D12-A [IgG3]) that detect circulating soluble egg antigen (CSEA) of Schistosoma mansoni were combined into one assay. This assay showed better performance than either of the two MAbs alone in detecting egg antigen, which was demonstrated with 80 urine samples from patients infected with Schistosoma mansoni from Zaire. The lower detection limit of the combined ELISA was 90 pg of the trichloroacetic acid-soluble fraction of soluble egg antigen (SEA-TCA) per milliliter. Thirty-two serum samples and 107 urine samples from uninfected Dutch individuals were negative when tested with the combined ELISA. This assay showed the same sensitivity (86.3%) with patients' urine samples as parallel testing with ELISA-5B1 and ELISA-4D12 (85%), while ELISA-5B1 and ELISA-4D12 showed sensitivities of 81.3% and 75%, respectively. The sensitivity of the combined ELISA with 51 serum samples was 84.3%, and three of five serum samples available from the seven patients with negative urine were positive for CSEA. The concentration of CSEA calculated from a four-parameters logistic curve for samples tested showed a correlation with egg output and serum circulating anodic antigen (P < 0.0001). Circulating soluble egg antigen in urine showed a significant decrease with an increase in age of the patients in relation to serum CSEA and egg output.

Age-dependent reduction of schistosome fecundity in Schistosoma haematobium but not Schistosoma mansoni infections in humans.

Understanding the dynamics of schistosome infections is problematic because direct measurements of worm burden are not possible. Hitherto, the relative intensity of infection has been estimated by the number of parasite eggs excreted. Egg excretion is assumed to have a consistent relationship with worm burden with duration of infection. We have tested this assumption in Schistosoma mansoni- and S. haematobium-infected populations by looking at the relationships between a circulating parasite antigen, egg excretion level, host age, and parasite density. The study was carried out in two populations because experimental models suggested that S. haematobium but not S. mansoni suffers immune-mediated reduction of fecundity. The results were consistent with this observation, showing that S. mansoni egg output remains stable irrespective of host age or infection intensity while S. haematobium has a substantially reduced egg production with host age. This information is fundamental to understanding the immunology and epidemiology of human schistosomiasis and thus practical approaches to disease control.

Levels of the schistosome circulating anodic and cathodic antigens in serum of schistosomiasis patients from Brazil.

Serum levels of 2 schistosome circulating antigens, the circulating anodic antigen (CAA) and the circulating cathodic antigen (CAA), were determined in persons infected with Schistosoma mansoni in Brazil. Sensitive monoclonal antibody-based enzyme-linked immunosorbent assays were used to measure levels of the 2 antigens. The study group consisted of 38 individuals with intestinal schistosomiasis, and 20 persons with the hepatosplenic form of the disease. Age and intensity of infection were comparable for the 2 groups. CAA was detected in 65.5% of all patients' sera and CCA was found in the serum of 82.8% of all patients. CAA levels correlated well with the egg output, as determined by duplicate Kato-Katz smears; CCA was significantly positively correlated with egg output in patients with intestinal schistosomiasis only. Whereas no significant difference was found between CAA titre in patients with intestinal schistosomiasis and those with the hepatosplenic form, a significantly higher CCA titre was found in patients with hepatosplenomegaly compared to patients with intestinal schistosomiasis.

Circulating anodic antigen levels in serum before and after chemotherapy with praziquantel in schistosomiasis mansoni.

The kinetics of serum levels of circulating anodic antigen (CAA) of Schistosoma mansoni were studied in patients with intestinal schistosomiasis before and after treatment with praziquantel. Day to day fluctuation in faecal egg excretion was compared with fluctuation in antigen level in 20 patients by serum and stool examination on 3 consecutive days before treatment. Antigen levels - calculated either as absorbance value of undiluted serum or as titre - showed less fluctuation than the number of eggs per gram of faeces determined by stool examinations based on single or duplicate 25 mg Kato smears. Compared with a placebo control group of 11 individuals, there was a significant reduction in CAA level in serum of 10 patients treated with praziquantel (40 mg/kg), 10 weeks after treatment. A similar decrease in serum CAA level was observed in a group of 46 patients treated with praziquantel, 6 weeks after treatment. In both groups, patients who remained seropositive after treatment still excreted eggs in their faeces. The kinetics of the antigen decrease were studied in more detail in 20 patients in hospital. Within 10 d after treatment with a double dose of 40 mg praziquantel per kg body weight, the antigen level fell to less than 10% of the original serum level, with a CAA half-life of approximately 2 d.

Detection of the schistosome circulating cathodic antigen by enzyme immunoassay using biotinylated monoclonal antibodies.

We have developed an enzyme immunoassay (ELISA) for the quantification of the schistosome circulating cathodic antigen (CCA), a glycoprotein associated with the syncitium lining the gut of the parasite. A mouse monoclonal antibody of IgG3 isotype was used as coating (antigen-capture) antibody, while a biotinylated mouse monoclonal IgM was used as second (antigen-detecting) antibody. Streptavidin-alkaline phosphatase was used as enzyme label. The lower detection limit of the assay was 1.0 ng of the trichloroacetic acid soluble fraction of adult worm antigen (AWA-TCA) per ml, which corresponds to approximately 0.2 ng CCA per ml. The ELISA showed a linear range from 1.0 to 62.5 ng AWA-TCA per ml. Serum and urine samples of 16 individuals infected with Schistosoma mansoni (egg counts ranging from 5 to 4820 eggs per gram of faeces) were tested in the assay. Antigen titres ranged from less than 4-8192. This assay represents a considerable advantage in diagnosis of Schistosoma infections as it allows the detection and quantification of CCA in serum and urine in even lightly infected individuals.

A simple technique to pretreat urine and serum samples for quantitation of schistosome circulating anodic and cathodic antigen.

For the detection of the circulating schistosome antigens CAA (circulating anodic antigen) and CCA (circulating cathodic antigen) in serum and urine samples of Schistosoma infected individuals, pretreatment of samples with trichloroacetic acid (TCA) is a standard procedure. In the present study several methods were evaluated in order to develop a more simple and rapid technique than the--especially for pretreatment of urine samples--laborious TCA technique. Optimal results were obtained with a method in which serum or urine samples were pretreated by a heat-incubation step (70 degrees C, 30 min) in an alkaline buffer (pH 9.6). In a comparison of the new technique with the TCA pretreatment, serum and urine samples of S. mansoni infected individuals from Zaire (n = 80) and of uninfected controls from The Netherlands (n = 208) were pretreated and assayed for CAA and CCA. Both pretreatment techniques showed similar sensitivities and specificities for CAA and CCA in serum, and CCA in urine. However, for the determination of CAA in urine the new technique performed significantly better, resulting in an increase of the sensitivity from 32 to 70% (titre determination).

Immunodiagnosis of schistosomiasis mansoni in a low endemic area in Surinam by determination of the circulating antigens CAA and CCA.

We evaluated the applicability of circulating antigen detection in serum and urine for the diagnosis of Schistosoma infections in a low endemic area. In total 389 individuals from Saramacca (Surinam) participated in the survey. Stool samples were examined using the Kato method, while circulating anodic antigen (CAA) and circulating cathodic antigen (CCA) were determined by highly specific monoclonal antibody-based ELISA's. Also schistosome specific IgM antibodies were measured by the indirect immunofluorescence assay, but the diagnostic performance of this test was found to be poor in this population. S. mansoni eggs were found in 29% of the examined cases, while CAA and CCA could be demonstrated in 23% and 17% of the serum samples and in 3% and 28% of the urine samples, respectively. Forty three percent of the study population was positive in at least one of these diagnostic assays, indicating that each individual test misses a substantial part of the subjects with an active infection. In most positive cases, intensities of infection were very low. As 204 individuals participated in all screening assays, diagnostic performance of each test was evaluated in this sub-population. The highest sensitivities were achieved with the urine-CCA assay and the parasitological examination, detecting 59 and 58 out of the 107 cases with an active infection, respectively. The serum-CAA assay detected 47 positive cases. Our results demonstrate that determination of circulating antigens, especially CCA in urine and CAA in serum, provides information additional to the parasitological examination, for the assessment of prevalence and intensity of Schistosoma infection in low endemic areas.

Presence of circulating anodic antigen in serum of Schistosoma intercalatum-infected patients from Gabon.

The presence of the schistosome circulating anodic antigen (CAA) in serum of Schistosoma intercalatum-infected patients from Gabon has been investigated using an enzyme-linked immunosorbent assay (ELISA). Blood samples were collected from 10 endemic controls, 29 patients which excreted viable S. intercalatum eggs in rectal mucosa and stool, six persons in which only non-viable eggs were found in rectal biopsy specimens and one person in which besides non-viable eggs a small number of viable eggs was found in the rectal biopsy specimen. CAA, a genus-specific antigen, could be demonstrated in 58.6% of the patients with S. intercalatum eggs in their stools. In comparison to S. mansoni infections, very light infections (0.6 eggs per gram faeces) could be detected by the ELISA. A strong correlation between parasite burden (eggs per gram faeces) and antigen-level (CAA-titer) was found (Spearman's rho = 0.65). Only one positive ELISA-results was found in the group with solely non-viable eggs in rectal tissue. As no false positive results were detected for the negative controls, the present results suggest, in accordance with results earlier obtained for schistosomiasis mansoni, that only in active S. intercalatum infections is antigen demonstrable.

Mixed Schistosoma haematobium and S. mansoni infection: effect of different treatments on the serum level of circulating anodic antigen (CAA).

In this study, levels of circulating anodic antigen (CAA) in serum were investigated after differential treatment of 160 Sudanese patients with mixed Schistosoma haematobium and S. mansoni infections. The patients were randomly divided into four groups, which were treated with metrifonate (two doses of 10 mg/kg bodyweight), oxamniquine (60 mg/kg), praziquantel (40 mg/kg), or a multivitamin preparation, respectively. Serum, stool and urine samples were taken prior to treatment as well as one month and five months after chemotherapy. Before chemotherapy CAA levels were similar in the four groups. Antigenemia remained unchanged in the control group. In patients treated with praziquantel or oxamniquine the concentration of CAA decreased to a similar extent. However, whereas in the praziquantel group absence of CAA was already observed one month after treatment, clearing of CAA from the circulation seemed to take longer in patients treated with oxamniquine. Treatment with metrifonate did not result in a reduction of the CAA titres.

Human IgE, IgG subclass, and IgM responses to worm and egg antigens in schistosomiasis haematobium: a 12-month study of reinfection in Cameroonian children.

Levels of IgE, IgM, and IgG subclasses against Schistosoma haematobium adult worm antigen (AWA) and soluble egg antigen (SEA) in a cohort of 148 S. haematobium-infected schoolchildren were determined before and up to 12 months after chemotherapy. Infection intensities were determined as concentrations of circulating anodic antigen (CAA) in serum. One month posttreatment, the antibody levels of all isotypes against AWA were increased, but 1 year after treatment they returned to pretreatment levels. CAA concentrations were positively associated with levels of IgG4 against AWA and SEA but not with levels of IgE. Age correlated negatively with CAA concentrations and positively with levels of IgE to AWA. The balance of anti-AWA IgG4 and IgE was significantly correlated to the CAA concentration, in particular in the older age group (11-13 years). This may suggest that protective immune mechanisms in S. haematobium infections become effective around the age of 12 years.

Immunodiagnosis of schistosomiasis patients in The Netherlands: comparison of antibody and antigen detection before and after chemotherapy.

We investigated in a retrospective study the application of an enzyme-linked immunosorbent assay (ELISA) for the quantitative detection of the schistosome circulating anodic antigen (CAA) to the diagnosis of schistosomiasis in our laboratory. CAA-titres were compared with IgG-titres as determined with an ELISA using soluble egg antigen (SEA) and IgM-titres as determined in an immuno fluorescence assay (IFA) on sections of Rossman's fixed schistosomes. Pre- and post-treatment serum samples of 182 individuals, suspected for schistosomiasis, were used for this study, including 100 control samples. CAA was detected in 20.7% of all patients who were positive in either IFA or SEA-ELISA or both. CAA-titres did not correlate with either IgM-titres or IgG-titres. No significant drop in IgG level was found after specific treatment (Wilcoxon's P = 0.35). In contrast, a significant fall in both IgM-titre (P less than 0.01), and CAA-level (P less than 0.01) was observed. All CAA-titres became negative after chemotherapy.

A model system for the study of antigen secretion by adult Schistosoma mansoni in vivo.

Schistosoma mansoni (6 weeks old) were surgically transferred from donor C57BI/6 mice to the hepatic portal veins of naive recipients of the same inbred strain. Between 70% and 100% of the parasites were alive 15 days later, and egg deposition was observed after transfer of worm pairs. The physiological status of the parasites was monitored by measuring the levels of a schistosome gut antigen, circulating anodic antigen (CAA), in the serum of the recipients. When only male worms were transferred, serum CAA levels increased slowly to a peak 9 days later, which was followed by a rapid decline. When worm pairs were transferred, there was an early peak in serum CAA levels followed by a gradual decline, but these levels were always higher than those recorded after male-only transfer; in two mice the pattern was similar to that observed following receipt of male worms. More CAA and eggs were produced after transfer of paired versus separated worms. It was concluded that although worm pairs can be successfully transferred, their physiological status may be sub-optimal. In contrast, male worms survive consistently well, and their transfer to a naive recipient provides a convenient model with which to study the release of antigens by schistosomes in vivo.

Monitoring the efficacy of different doses of praziquantel by quantification of circulating antigens in serum and urine of schistosomiasis patients.

We evaluated the quantitation of two schistosome circulating antigens in serum and urine as a tool for the assessment of the efficacy of praziquantel dosage regimens (40 versus 60 mg/kgbw). In addition we compared the efficacy of two different brands of praziquantel (Biltricide and Distocide), given at the same dosage (40 mg/kgbw). Thirty five Egyptian hospitalized schistosomiasis mansoni patients participated in this study. Thirteen patients (Group 1) received 60 mg/kgbw Biltricide, administered in 3 oral doses of 20 mg in one day; 22 individuals (Group 2) were treated with 40 mg/kgbw (12 Biltricide, 10 Distocide), given in one oral dose. Circulating anodic antigen (CAA) and circulating cathodic antigen (CCA) were quantitated by monoclonal antibody-based ELISA's before, and 1, 3 and 6 weeks after chemotherapy. Before treatment, all patients were positive for at least one of the circulating antigen assays. Three to six weeks after treatment significantly more patients were found to be negative in Group 1 compared to Group 2 (X2 = 7.13, P = 0.008, n = 35). Also the levels of CCA and CAA in serum and of CCA in urine were found to be significantly higher in Group 2 (Mann-Whitney U < 85, P < 0.05, n = 35). These results were confirmed by parasitological data. No differences were found between treatment with Biltricide or Distocide.(ABSTRACT TRUNCATED AT 250 WORDS)

Circulating anodic and cathodic antigen in serum and urine from Schistosoma haematobium-infected Cameroonian children receiving praziquantel: a longitudinal study.

A cohort of 148 Cameroonian children infected with Schistosoma haematobium was followed before praziquantel therapy and 1, 2, 3, 5, and 12 months thereafter. Egg output, the reagent strip index (RSI, a pathological marker), and circulating anodic antigen (CAA) and circulating cathodic antigen (CCA) in serum and urine were quantified. At enrollment, the median level of egg output was 365/10 mL of urine; 97% of children had a positive RSI; CAA was detected in serum from 76% of children and in urine from 64%; and CCA was detected in serum from 55% of children and in urine from 87%. Two months after chemotherapy, egg output and RSI had decreased significantly; reinfection later developed in parallel with increases in the serum and urine concentrations of CAA and the urine concentrations of CCA. The measurement of CAA and CCA is useful for diagnosis, evaluation of disease severity, and follow-up of chemotherapy in individuals infected with S. haematobium.