Diagnosis of invasive amebiasis by enzyme-linked immunosorbent assay of saliva to detect amebic lectin antigen and anti-lectin immunoglobulin G antibodies.

Saliva from subjects with amebic liver abscess (ALA), acute amebic colitis, asymptomatic infection with Entamoeba histolytica or Entamoeba dispar, and uninfected controls was tested by enzyme-linked immunosorbent assay (ELISA) for the presence of E. histolytica galactose-inhibitable lectin antigen and salivary immunoglobulin (IgG) antibodies to a recombinant cysteine-rich lectin-derived protein (LC3). Salivary lectin antigen was found in 65.8% of subjects with acute colitis, compared to 22.2% of those convalescent from ALA, 10.0% with asymptomatic E. histolytica infection, 9.8% with E. dispar infection, and 2.6% of controls (subjects from the United States and study patients with nonamebic diarrhea) (P < 0.001 for each compared to values for subjects with colitis). Salivary anti-LC3 IgG antibodies were found in 92% of ALA patients regardless of duration of illness and in 83.3% of colitis patients who were symptomatic for at least 7 days (P < 0.001 compared to other study groups). Serum anti-LC3 IgG antibodies were detected in 56.3% of subjects with acute colitis, 100% of subjects with ALA or prolonged colitis, 45% of subjects with asymptomatic E. histolytica infection, 32.3% of subjects with E. dispar infection, and 23.4% of diarrhea controls. In comparison to ELISA for serum anti-LC3 IgG antibodies, the salivary lectin antigen assay is a more sensitive and specific test for acute amebic colitis. Detection of salivary anti-LC3 IgG antibodies by ELISA is an effective means for the diagnosis of ALA and prolonged cases of amebic colitis.

Comparison of antigen-capture ELISA to stool-culture methods for the detection of asymptomatic Entamoeba species infection in Kafer Daoud, Egypt.

We performed a prospective field study in the village of Kafer Daoud in Menofia, Egypt to compare the fecal culture method with enzyme linked immuno assay (ELISA) for detection of 170 kDa lectin antigen in feces for diagnosis of asymptomatic Entamoeba histolytica and Entamoeba dispar infection. All subjects with E. histolytica or E. dispar infection detected by culture also had positive ELISA for amebic antigen in their feces and an additional 57 Entameoba infections missed by culture were detected by ELISA (P < 0.001 compared to culture). The presence of fecal anti-lectin IgA antibodies and serum anti-LC3 (recombinant cysteine-rich lectin protein) IgG antibodies were positive predictors for E. histolytica infection (P < 0.03). Of interest, infection with Trichomonas hominis but not Blastocystis hominis was positively associated with E. histolytica infection (P < 0.05). In conclusion, ELISA for detection of fecal lectin antigen is a more sensitive method than fecal culture for detecting asymptomatic E. histolytica infection.

Serum IgM antibody response to the galactose-inhibitable adherence lectin of Entameoba histolytica.

An ELISA for detection of serum IgM antibodies to the galactose-inhibitable adherence lectin of Entamoeba histolytica revealed that 2.8% of uninfected controls, 0.0% of controls infected with other parasites, 13.4% of asymptomatic amebic infections, 55% of colitis patients, and 77% of amebic liver abscess patients from Cairo, Egypt and Durban, South Africa had serum anti-lectin IgM antibodies. Of acute amebic colitis patients with symptoms for less than one week, only 6% possessed serum IgG anti-lectin antibodies, yet 45% had serum IgM antibodies to the amebic lectin. This compares with 65% of sera in acute colitis patients positive for lectin antigen as determined by ELISA with anti-lectin monoclonal antibodies. In conclusion, an ELISA for serum anti-lectin IgM antibodies appears to have greater clinical utility in the setting of acute amebic colitis than an ELISA for anti-lectin IgG antibodies, but is no more sensitive than an ELISA for detection of lectin antigen in sera.

Protection of gerbils from amebic liver abscess by immunization with recombinant Entamoeba histolytica 29-kilodalton antigen.

The goal of our study was to obtain a highly conserved Entamoeba histolytica recombinant antigen for study as a subunit amebiasis vaccine. We screened a Uni-Zap cDNA library of E. histolytica (strain HM1:IMSS) with human immune sera and isolated a dominant 804-bp cDNA clone. A 33-kDa fusion protein expressed from the cDNA clone was determined by monoclonal antibody binding, DNA hybridization, and nucleotide sequence to be the complete E. histolytica 29-kDa antigen. Serum antibodies to the recombinant protein were detected by enzyme-linked immunosorbent assay in 80% of subjects from Egypt and South Africa with amebic liver abscess. Similar results were found with the native 29-kDa protein. Native and recombinant 29-kDa antigens induced proliferation of lymphocytes harvested from patients with amebic liver abscess (P < 0.01 compared with controls). Intraperitoneal immunization of gerbils with the recombinant fusion protein (10 micrograms) with Titermax adjuvant elicited an antigen-specific serum immunoglobulin G antibody response and was partially protective (54%) against intrahepatic challenge with 5 x 10(5) virulent axenic trophozoites (strain HM1:IMSS). In summary, the recombinant form of the E. histolytica 29-kDa antigen demonstrated serologic specificity for amebic liver abscess, exhibited conserved T-cell epitopes, and was effective as a subunit vaccine in an experimental animal model of amebic liver abscess.

Differentiation of pathogenic Entamoeba histolytica infections from nonpathogenic infections by detection of galactose-inhibitable adherence protein antigen in sera and feces.

We determined whether epitope-specific monoclonal antibodies to the galactose-inhibitable adherence protein (GIAP) of Entamoeba histolytica could be used in an enzyme-linked immunosorbent assay (ELISA) to detect antigen in serum and feces and differentiate between nonpathogenic zymodemes and the potentially invasive pathogenic organisms that require treatment. Overall, 57% of subjects from Cairo, Egypt, with symptomatic intestinal amebiasis and 42% with asymptomatic infection possessed GIAP antigen in their sera, whereas 4% of uninfected controls or subjects with other parasitic infections possessed GIAP antigen in their sera (P < 0.001). In subjects from Durban, South Africa, only 6% of uninfected controls or those with nonpathogenic E. histolytica infection were positive for GIAP in serum, whereas 3 of 4 with asymptomatic pathogenic intestinal infection and 75% with amebic liver abscess were positive for GIAP in serum. Fifteen stool samples from patients with intestinal amebiasis were available for study; all had a positive ELISA result for fecal GIAP antigen. Epitope-specific monoclonal antibodies identified 8 of 15 subjects with fecal antigen from pathogenic strains. Seven of those eight subjects had adherence protein antigen in their sera, whereas none of seven with apparent nonpathogenic E. histolytica infection had adherence protein antigen in their sera. In summary, we were able to detect E. histolytica adherence protein antigen directly in serum and fecal samples by ELISA. The presence of amebic antigen in serum demonstrated 94% specificity for pathogenic E. histolytica infection, and amebic antigen is present during asymptomatic intestinal infection. In conjunction with antibody detection, this method should be very useful in the diagnosis and management of intestinal amebiasis.

Use of an enzyme-linked immunosorbent assay to detect anti-adherence protein antibodies in sera of patients with invasive amebiasis in Cairo, Egypt.

We used enzyme-linked immunosorbent assay (ELISA) to detect IgG antibodies to the Entamoeba histolytica galactose-inhibitable adherence protein in the sera of 50 uninfected controls, 50 cases with asymptomatic cyst passage, 100 patients with amebic colitis, and six patients with amebic liver abscess from Cairo, Egypt, and in 50 healthy controls from the United States. When the mean + 3 SD value above that of the controls from the United States was used as a criterion for a positive ELISA result, 100% of those with invasive amebiasis, 80% of those with asymptomatic infection, and 64% of the Egyptian controls had anti-adherence protein antibodies. However, when the mean + 2 SD value of Egyptian control sera (optical density = 0.094) was used as the criterion for positivity, 33 (89%) of 37 sera from individuals with invasive amebiasis having symptoms for at least one week were antibody positive, in contrast to only 12% of asymptomatic cyst passers (P < 0.01). In a highly endemic area such as Cairo, Egypt, detection of serum anti-adherence protein antibodies by ELISA may have greatest diagnostic use in patients with symptomatic invasive amebiasis of greater than one week duration.