Antibodies against glucan, chitin, and Saccharomyces cerevisiae mannan as new biomarkers of Candida albicans infection that complement tests based on C. albicans mannan

Abstract

Antibodies against Saccharomyces cerevisiae mannan (ASCA) and antibodies against synthetic disaccharide fragments of glucans (ALCA) and chitin (ACCA) are biomarkers of Crohn's disease (CD). We previously showed that Candida albicans infection generates ASCA. Here, we explored ALCA and ACCA as possible biomarkers of invasive C. albicans infection (ICI). ASCA, ALCA, ACCA, and Candida mannan antigen and antibody detection tests were performed on 69 sera obtained sequentially from 18 patients with ICIs proven by blood culture, 59 sera from CD patients, 47 sera from hospitalized subjects colonized by Candida species (CZ), and 131 sera from healthy controls (HC). ASCA, ALCA, and ACCA levels in CD and ICI patients were significantly different from those in CZ and HC subjects (P<0.0001). In ICI patients, these levels increased as infection developed. Using ASCA, ALCA, ACCA, and Platelia Candida tests, 100% of ICIs were detected, with the kinetics of the antibody response depending on the patient during the time course of infection. A large number of sera presented with more than three positive tests. This is the first evidence that the detection of antibodies against chitin and glucans has diagnostic value in fungal infections and that these tests can complement more specific tests. Future trials are necessary to assess the value of these tests in multiparametric analysis, as well as their pathophysiological relevance.

FIG. 1.

Distribution of ASCA, ALCA, and ACCA in healthy controls, patients with Crohn's disease, ICU patients with one or two body sites colonized by Candida species, and patients with invasive candidiasis. Comparison of the values for the different groups of patients was performed using the Mann-Whitney U test (P values). ASCA, ALCA, and ACCA results were expressed in arbitrary units (AU) (see Materials and Methods). For ASCA, a highly significant difference was observed for HC versus CD patients (P < 0.0001) and HC versus ICI patients, while the difference between CD versus ICI patients was not statistically significant (NS). For ALCA, a highly significant difference was observed for HC versus CD patients (P < 0.0001) and HC versus ICI patients (P < 0.0001), while no difference was observed for CD versus ICI patients. For ACCA, similar results were observed for HC versus CD patients (P = 0.001) and HC versus ICI patients (P < 0.0001). In contrast to ASCA and ALCA, a significant difference in ACCA was observed between ICI and CD patients (P = 0.05). The same trend was observed for CZ patients, and no difference was observed for HC versus CZ patients. The chemical structures of the antigens are presented to the right of each graph; for ALCA and ACCA, synthetic oligosaccharides are coated on the wells of the ELISA plates; ASCA* antigen is a natural antigen which comprises a repertoire of oligomannose epitopes, among these we have represented the major epitope supporting the humoral response in CD patients (39, 52), since this synthetic analog was shown to specifically adsorb antibodies generated during C. albicans infection (43).

FIG. 2.

(a) Development of anti-C. albicans mannan antibodies (Platelia Ab [Plat. Ab]), ASCA, ALCA, and ACCA in New Zealand White rabbits following intravenous inoculation of live C. albicans strain VW32. Results are expressed as mean ODs plus standard errors (error bars). D0, day 0. (b) Murine MAb 2G8 (gray bars) and MAb EB-CA1 (black bars) reactivities were determined by ELISA with laminaribioside, a synthetic analog of β-1,3 glucan involved in the ALCA test.

FIG. 3.

Sixty-nine serum samples from 18 patients with invasive Candida infection (ICI) were screened for the presence of ASCA (a), ALCA (b), ACCA (c), and Platelia Ab (Plat. Ab) (d) as described in Materials and Methods. The antibody levels in arbitrary units (AU) (see Materials and Methods) are plotted according to the date of serum sampling (day 0 indicates the date of mycological isolation of C. albicans from blood). The horizontal line indicates the cutoff values used to define positive and negative results. The vertical line indicates day 0. Antibody values (mean titers plus standard errors [error bars]) for each test are also presented as histograms (panels a2, b2, c2, and d2) by classifying sera into four groups as follows: group 1 (G1) for sera taken during the period from day −25 to day −1, group 2 (G2) for sera taken from day 0 to day +15; group 3 for sera taken from day +16 to day +40; and group 4 for sera taken from day +41 to day +154.

FIG. 4.

Examples of the kinetics of ASCA, ALCA, ACCA, and Platelia Candida Ag and Ab tests in patients with proven invasive candidiasis. For both patients, patient 14 (a) and patient 6 (b), a gradual decrease in ASCA, ALCA, and ACCA was observed during the period preceding positive blood cultures to reach a minimum on day 0. After the candidemic episode, an overall increase could be observed for most antibody markers during the proceeding weeks. For each y axis, the symbols on the axis indicate the cutoff values for serological tests. ASCA, ALCA, ACCA, and Platelia Candida Ab titers are given in arbitrary units (AU) (see Materials and Methods).