Immunodiagnosis of Fasciola hepatica infection (fascioliasis) in a human population in the Bolivian Altiplano using purified cathepsin L cysteine proteinase.

Abstract

Cathepsin L1 (CL1), an immunogenic cysteine proteinase secreted by juvenile and adult Fasciola hepatica, was assessed for its potential as a diagnostic agent for the serologic detection of human fascioliasis. Using ELISAs, we compared the ability of liver fluke homogenates (LFH), excretory/secretory (ES) products, and CL1 to discriminate between seropositive (infected) and seronegative (noninfected) individuals within a population of 95 patients from the Bolivian Altiplano. A high prevalence of human fascioliasis has been reported in this region. The division between the seropositive and seronegative individuals was poorly defined when LFH was used as the antigen. A greater discrimination between these populations was achieved with both ES and CL1. A K-means cluster analysis using the combined ES and CL1 ELISA data identified a cluster of seropositive individuals. Cathepsin L1 detected a subset (20) of these seropositive individuals while ES detected all 26; however, ES detected nine additional individuals that were in the seronegative cluster. The ratio of the mean absorbance readings between seropositive and seronegative individuals was markedly improved by using conjugated second antibodies to IgG4, the predominant isotype elicited by infection. In these IgG4-ELISAs, CL1 again identified fewer individuals as seropositive than did ES, but improved the discrimination between the seropositive and seronegative individuals and thus provided a more conclusive diagnosis. Sera obtained from patients infected with schistosomiasis mansoni, cysticercosis, hydatidosis, and Chagas' disease were negative in these assays, which demonstrated the specificity of the IgG4-ELISA for detecting fascioliasis. Twenty of the 95 patients (21%) were seropositive for fascioliasis by the CL1 IgG4-ELISA, confirming the earlier reports of the high prevalence of disease in this region. A standardized diagnostic test for human fascioliasis, based on an ELISA that detects IgG4 responses to CL1, could be available to all diagnostic centers if sufficient quantities of recombinant CL1 can be produced.

Short report: Diagnosis of human fascioliasis: detection of anti-cathepsin L antibodies in blood samples collected on filter paper.

“…The assay detects serum IgG4 antibodies reactive with Fasciola hepatica cathepsin L1. 1 Using this diagnostic assay, we confirmed earlier reports 2-4 of a high incidence (22% of the population analyzed) of human fascioliasis in Calasaya, a village in the northern Altiplano of Bolivia. Further studies to examine the extent of this disease in the Altiplano region were hampered by the reluctance of the indigenous Aymaran population to provide blood by venipuncture.…”

“…We therefore analyzed the sera of all coprologically negative individuals from Cutusuma and Chijipata by the IgG4-ELISA. We then used a statistical method, hierarchical agglomerative cluster analysis, 1,6 to identify subpopulations that may represent individuals that were exposed or not exposed to fasciolosis (Figure 1b). This analysis separated the population into a low absorbance group, which represents the nonexposed individuals, and a high absorbance group, which represents the exposed individuals.…”

Short report: Immunodiagnosis of human fascioliasis using recombinant Fasciola hepatica cathepsin L1 cysteine proteinase.

“…5-8 Our laboratory recently developed a simple ELISA for the detection of serum IgG4 antibodies reactive with F. hepatica cathepsin L1 (CL1) proteinase. 7 The use of purified CL1 antigen in this ELISA provided a more specific immunodiagnosis of human fascioliasis than tests that use parasite somatic or excretory/secretory (ES) antigen.We have also recently reported the expression of functionally active recombinant F. hepatica CL1 in the yeast Saccharomyces cerevisiae. 9 In the present study, we examined the potential of this recombinant protein as a diagnostic reagent for human fascioliasis in the IgG4-ELISA and compared the performance of the antigen with native CL1 antigen.…”

“…5-8 Our laboratory recently developed a simple ELISA for the detection of serum IgG4 antibodies reactive with F. hepatica cathepsin L1 (CL1) proteinase. 7 The use of purified CL1 antigen in this ELISA provided a more specific immunodiagnosis of human fascioliasis than tests that use parasite somatic or excretory/secretory (ES) antigen.…”

“…3,4 The disease is particularly prevalent among the Aymaran people of the northern Altiplano region of Bolivia, with incidences of up to 60% being reported. [5][6][7][8] Our laboratory recently developed a simple ELISA for the detection of serum IgG4 antibodies reactive with F. hepatica cathepsin L1 (CL1) proteinase. 7 The use of purified CL1 antigen in this ELISA provided a more specific immunodiagnosis of human fascioliasis than tests that use parasite somatic or excretory/secretory (ES) antigen.…”