Abstract
Anti-β-glucan antibodies elicited by a laminarin-conjugate vaccine confer cross-protection to mice challenged with major fungal pathogens such as Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans. To gain insights into protective β-glucan epitope(s) and protection mechanisms, we studied two anti-β-glucan monoclonal antibodies (mAb) with identical complementarity-determining regions but different isotypes (mAb 2G8, IgG2b and mAb 1E12, IgM). C. albicans, the most relevant fungal pathogen for humans, was used as a model.Both mAbs bound to fungal cell surface and to the β1,3-β1,6 glucan of the fungal cell wall skeleton, as shown by immunofluorescence, electron-microscopy and ELISA. They were also equally unable to opsonize fungal cells in a J774 macrophage phagocytosis and killing assay. However, only the IgG2b conferred substantial protection against mucosal and systemic candidiasis in passive vaccination experiments in rodents. Competition ELISA and microarray analyses using sequence-defined glucan oligosaccharides showed that the protective IgG2b selectively bound to β1,3-linked (laminarin-like) glucose sequences whereas the non-protective IgM bound to β1,6- and β1,4-linked glucose sequences in addition to β1,3-linked ones. Only the protective IgG2b recognized heterogeneous, polydisperse high molecular weight cell wall and secretory components of the fungus, two of which were identified as the GPI-anchored cell wall proteins Als3 and Hyr1. In addition, only the IgG2b inhibited in vitro two critical virulence attributes of the fungus, hyphal growth and adherence to human epithelial cells.Our study demonstrates that the isotype of anti-β-glucan antibodies may affect details of the β-glucan epitopes recognized, and this may be associated with a differing ability to inhibit virulence attributes of the fungus and confer protection in vivo. Our data also suggest that the anti-virulence properties of the IgG2b mAb may be linked to its capacity to recognize β-glucan epitope(s) on some cell wall components that exert critical functions in fungal cell wall structure and adherence to host cells.
Highlights
Diseases caused by fungi are increasingly impacting on the health of populations, constituting a large fraction of health care-associated infections
In the search for protective anti-b-glucan antibodies analogous to those elicited by the Lam-CRM vaccine, we generated a number of anti-b-glucan murine monoclonal antibodies
The deduced amino acid sequences of variable regions of the two monoclonal antibodies (mAb) (Table 1) showed a complete identity of their three heavy and light chain Complementarity Determining Regions (CDRs). Both antibodies were found to bind to C. albicans germ-tubes and A. fumigatus hyphae (Figure 1, panel A, a– b; e–f)
Summary
Diseases caused by fungi are increasingly impacting on the health of populations, constituting a large fraction of health care-associated infections. Subject categories at high risk of fungal infection are cancer patients under immunosuppressive chemotherapy, subjects undergoing major surgery and critically-ill patients under supportive ventilation and bearing central venous and urinary catheters [1,2,3] Both diagnosis and antifungal therapy are of limited effectiveness in these patients, resulting into treatment failures and associated mortality [2,4,5]. The spectrum of fungal pathogens has enlarged to include yeasts and moulds that are refractory to most antifungals, posing remarkable challenges to infection control measures [1,3,6] In this context, it is much hoped that immuno-prophylactic or therapeutic treatments will be developed to drastically reduce the incidence of fungal infections and resulting mortality. Development of clinically useful antifungal antibodies may profit from the on-going outstanding advances in recombinant DNA technologies and protein engineering
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