Protection against candidiasis by an immunoglobulin G3 (IgG3) monoclonal antibody specific for the same mannotriose as an IgM protective antibody

Abstract

We previously reported that a liposome-mannan vaccine (L-mann) of Candida albicans induces production of mouse antibodies that protect against disseminated candidiasis and vaginal infection. Immunoglobulin M (IgM) monoclonal antibody (MAb) B6.1, specific for a C. albicans cell surface beta-1,2-mannotriose, protects mice against both infections. Another IgM MAb, termed B6, which is specific for a different cell surface mannan epitope, does not protect against disseminated candidiasis. The B6.1 epitope is displayed homogeneously over the entire cell surface, compared to a patchy distribution of the B6 epitope. To determine if protection is restricted to an IgM class of antibody, we tested an IgG antibody. MAb C3.1 was obtained from L-mann-immunized mice. By results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, capture enzyme-linked immunosorbent assay, and immunodiffusion tests, MAb C3.1 is an IgG3 isotype. By epitope inhibition assays, we determined that MAb C3.1 is specific for same mannotriose as MAb B6. 1. As expected by the results of the inhibition assays, immunofluorescence microscopy showed that the C3.1 epitope is distributed on the yeast cell surface in a pattern identical to that of the B6.1 epitope. Kidney CFU and mean survival times of infected mice pretreated with MAb C3.1 indicated that the antibody enhanced resistance of mice against disseminated candidiasis. Mice in pseudoestrus that were given MAb C3.1 prior to vaginal infection developed fewer vaginal Candida CFU than control animals that received buffered saline instead of the antibody. The finding that an IgG3 antibody is protective is consistent with our hypothesis that epitope specificity and complement activation are related to the ability of an antibody to protect against candidiasis.

FIG. 1

The C3.1 epitope is uniformly distributed on the cell surface of yeast forms of C. albicans. Hydrophilic stationary-phase yeast cells were reacted with MAb C3.1, washed, and counterreacted with fluorescence-labeled anti-mouse IgG. The C3.1 epitope is located over the entire cell surface (A). The same field was photographed under phase-contrast microscopy (B). The C3.1 epitope distribution displays the same homogeneous pattern over the cell surface as the epitope recognized by the IgM protective antibody, MAb IgM B6.1. These results provide further evidence that the IgG3 antibody is specific for the same oligomannoside as the protective IgM. Note that some yeast cells (arrows) and all of the blastoconidia (arrowheads) were nonreactive with MAb C3.1. The importance of nonreactive cells in pathogenesis and protection experiments is unknown. Bar, 10 μM.

FIG. 2

MAb C3.1 transfers protection against disseminated candidiasis. (A) Mice given unheated MAb C3.1 (C3.1), H-C3.1, A-C3.1, or DPBS (buffer diluent) i.p.; (B) mice given MAb C3.1, MAb B6.1, or DPBS as a control. In both panels, the animals were challenged i.v. with 5 × 10⁵ viable yeast cells, and susceptibility to disseminated disease was assessed by determining kidney CFU (A) and by survival curves (B). (A) Mice that received the unheated or heated MAb C3.1 had 86 and 88% fewer CFU, respectively, than mice that received DPBS (P < 0.001). Mice that were given the absorbed serum developed almost the same number of CFU as mice that received DPBS (control). Bars show standard errors. (B) Mice given MAb C3.1 had survival times similar to those of mice given MAb B6.1. Their mean survival times were significantly longer than for animals given DPBS (P < 0.05).

FIG. 3

MAb C3.1 has a prophylactic effect against Candida vaginal infection. Pseudoestrus mice were given MAb C3.1 i.p. (A) or i.vg. (B) before an i.vg. challenge with 5 × 10⁵ yeast cells. Vaginal CFU counts were compared with those from animals given unheated C3.1, H-C3.1, A-C3.1, or DPBS (diluent). Mice that received the unheated or heated C3.1 developed 60 and 49% fewer CFU, respectively, than control mice that received DPBS. Mice that received A-C3.1 or DPBS developed similar numbers of CFU (A). Mice that received the unheated or heated MAb C3.1 developed approximately 86% fewer CFU than DPBS control mice, whereas CFU numbers in mice given A-C3.1 were similar to those in the DPBS control mice (B). In both panels, significant differences were found between mice that received either the unheated MAb C3.1 or heated MAb C3.1 and DPBS controls (P < 0.05). Bars show standard errors.