Immunity to Entamoeba species intestinal
infection is associated with the presence of intestinal
IgA antibodies against the parasite's
galactose-inhibitable adherence
lectin. We determined the
epitope specificity of serum and intestinal antilectin
IgA antibodies by
enzyme-linked
immunosorbent assay using overlapping fragments of a recombinant portion of the
lectin heavy subunit, designated LC3. These findings were correlated with the effects of
epitope-specific murine antilectin
immunoglobulin A (
IgA)
monoclonal antibodies (MAbs) on amebic in vitro
galactose-specific adherence. LC3 is a highly antigenic and immunogenic
cysteine-rich
protein (
amino acids [aa] 758 to 1150) that includes the
lectin's
carbohydrate binding domain. The study subjects, from Durban, South Africa, were recently cured of
amebic liver abscess (ALA) with or without concurrent Entamoeba histolytica intestinal
infection or were
infection free 1 year after cure. We also studied seropositive subjects that were infected with E. histolytica, disease free, and asymptomatic. Serum anti-LC3
IgA antibodies from all study groups exclusively recognized the third (aa 868 to 944) and the seventh (aa 1114 to 1134) LC3
epitopes regardless of clinical status;
epitope 6 (aa 1070 to 1114) was also recognized by serum anti-LC3
IgG antibodies. However,
IgG antibody recognition of
epitope 6 but not 3 or 7 was lost 1 year following cure of ALA. We produced 14 murine anti-LC3
IgA MAbs which collectively recognized five of the seven LC3
epitopes. The majority of the murine MAbs recognized the first
epitope (aa 758 to 826), which was not recognized by human
IgA antibodies. Interestingly, adherence of E. histolytica trophozoites to CHO cells was inhibited by MAbs against
epitopes 1, 3, 4 (aa 944 to 987), and 6 (P < 0.01). The LC3
epitopes recognized by human
IgA antibodies (3 and 7) were further characterized by use of overlapping synthetic
peptides. We identified four
peptides (aa 891 to 903, 918 to 936, 1114 to 1134, and 1128 to 1150) that in linear or cyclized form were recognized by pooled intestinal
IgA antibodies and serum
IgG antibodies from subjects with ALA and asymptomatic, seropositive infected subjects. This study identifies the
lectin epitopes to be studied in an
amebiasis subunit vaccine designed to elicit mucosal immunity mimicking that of humans cured of ALA.