Interaction of Candida albicans Cell Wall Als3 Protein with Streptococcus gordonii SspB Adhesin Promotes Development of Mixed-Species Communities

Candida albicans colonizes human mucosa and prosthetic surfaces associated with artificial joints, catheters, and dentures. In the oral cavity, C. albicans coexists with numerous bacterial species, and evidence suggests that bacteria may modulate fungal growth and biofilm formation. Streptococcus gordonii is found on most oral cavity surfaces and interacts with C. albicans to promote hyphal and biofilm formation. In this study, we investigated the role of the hyphal-wall protein Als3p in interactions of C. albicans with S. gordonii. Utilizing an ALS3 deletion mutant strain, it was shown that cells were not affected in initial adherence to the salivary pellicle or in hyphal formation in the planktonic phase. However, the Als3⁻ mutant was unable to form biofilms on the salivary pellicle or deposited S. gordonii DL1 wild-type cells, and after initial adherence, als3Δ/als3Δ (ΔALS3) cells became detached concomitant with hyphal formation. In coaggregation assays, S. gordonii cells attached to, and accumulated around, hyphae formed by C. albicans wild-type cells. However, streptococci failed to attach to hyphae produced by the ΔALS3 mutant. Saccharomyces cerevisiae S150-2B cells expressing Als3p, but not control cells, supported binding of S. gordonii DL1. However, S. gordonii Δ(sspA sspB) cells deficient in production of the surface protein adhesins SspA and SspB showed >50% reduced levels of binding to S. cerevisiae expressing Als3p. Lactococcus lactis cells expressing SspB bound avidly to S. cerevisiae expressing Als3p, but not to S150-2B wild-type cells. These results show that recognition of C. albicans by S. gordonii involves Als3 protein-SspB protein interaction, defining a novel mechanism in fungal-bacterial communication.Candida species are the fourth most common causative agents of nosocomial bloodstream infections (2, 47, 54). Crude mortality rates for Candida infections exceed 50% (10, 52), and attributable mortality rates vary between 5 and 48% (3, 10, 13). Candida albicans accounts for 62% of invasive candidiasis infections (46, 47) and is commonly isolated from the oral cavity, gastrointestinal tract, and vagina. The oral carriage rate of C. albicans in healthy subjects ranges from 25 to 60% (28, 42, 48). In the oral cavity, there are estimated to be approximately 700 different species of microorganisms present (45). C. albicans is able to interact physically, by coaggregation, or chemically, through small-molecule signaling, with some of these other microorganisms (1, 18, 20, 29, 33). Interactions of C. albicans with bacteria may be antagonistic, e.g., with Pseudomonas aeruginosa (20), or synergistic, e.g., with Streptococcus gordonii (1), resulting in the formation of diverse polymicrobial communities.Streptococcus gordonii is a primary colonizer of the oral cavity and may be isolated from mucosal or hard surfaces present there (17, 41). It has previously been shown that S. gordonii, and other viridans streptococci, can coaggregate with C. albicans cells both in vitro and in vivo (21, 29, 57). The interactions between oral streptococci and C. albicans are recognized as contributing to formation of enhanced biofilms (1), which may occur on dentures, leading to denture stomatitis (42). Oral streptococci express a range of cell surface polypeptides, many of which act as adhesins to promote colonization (31, 38). The antigen (Ag) I/II family of polypeptides are cell wall-anchored proteins produced by most indigenous species of oral streptococci (4). These adhesins have been shown to bind a wide range of host cell proteins, including fibronectin (49) and salivary agglutinin gp-340 (5, 12, 27). In addition, the Ag I/II family polypeptide SspB from S. gordonii has been shown to interact directly with other microorganisms, including Actinomyces naeslundii (27), Porphyromonas gingivalis (11), and C. albicans (1, 22). It is thus proposed that oral streptococci may promote colonization by these other microorganisms by providing alternative surfaces to adhere to (30) and possibly metabolic benefits (25).Candida albicans is a pleomorphic fungus, with the two most commonly identified morphologies being yeast cells and hyphae. Hyphal-filament formation may be induced by many factors, including pH, serum, temperature, nutrient availability, and diffusible cell signaling molecules (53). In a mixed-species biofilm model, S. gordonii enhances hyphal formation, and there is evidence that this may be mediated, at least in part, by soluble factors released by streptococci (1). Within mixed-species biofilms of S. gordonii and C. albicans, streptococci were found associated with yeast cells, pseudohyphae, and hyphae, but preferentially with hyphal filaments (1).The hyphal cell wall comprises a mixture of chitin, β-1,3 glucans, and β-1,6 glucans, as well as a vast array of proteins (7). One of the major families of C. albicans adhesins is the ALS (agglutinin-like sequence) group of cell wall glycoproteins (24). The family comprises 8 members, several of which have adhesive functions involved in host-pathogen interactions (24). One of these adhesins, Als3p, is a hypha-specific protein (9, 23) and has been shown to be required for mature-biofilm formation, binding extracellular matrix, adhesion to host cells, and internalization of C. albicans by endothelial cells (24, 50, 56). There is also evidence that the Als5 protein is involved in recognition of S. gordonii by C. albicans (32).In this study, we investigated the role of hypha-specific Als3p in early-stage biofilm formation and in intergeneric interactions of C. albicans with S. gordonii. The results suggest that Als3p interacts directly with SspB on the surface of S. gordonii, a binding event that may then enable additional concerted adhesin-receptor interactions to become established.