Bacterial keratitis is a sight-threatening infectious disease of the cornea caused by different types of bacteria, including Staphylococcus aureus, Streptococcus pneumoniae and Pseudomonas aeruginosa (3). One of the first steps during bacterial infection is the adhesion of the pathogen to glycoproteins and glycolipids on the epithelial cell glycocalyx through specific glycan recognition mechanisms (4, 5). Notably, this adhesive interaction does not take place unless the epithelia are damaged (6). Mucins stand out among the multiple protective components of the healthy glycocalyx because of their ability to limit infectious disease while accommodating resident microbiota (7). Transmembrane mucins have large and rigid extracellular domains that extend 500 nm or more above the cell surface. They can prevent microbial colonization by several mechanisms that include forming a physical barrier, acting as adhesion decoys and, in certain cases, exposing specific glycans that attenuate pathogen virulence.

The presence of the transmembrane mucin MUC16 at the ocular surface is a major restrain to the passage of bacteria. Reports using a human cell culture model of stratified corneal epithelium have evidenced that MUC16 and mucin O-glycans protect the epithelial surface against S. aureus adhesion (8, 9). It has been established that transmembrane mucins in cornea play a crucial role in the early response against pathogens by suppressing Toll-like receptor signaling and the expression of the proinflammatory cytokines (10). Therefore, in the absence of pre-existing defects or wounding, non-opportunistic bacteria must rely on the enzymatic removal of transmembrane mucins to access epithelial cells and cause disease. This is the case with epidemic disease-causing S. pneumoniae species, which secrete a metalloproteinase, ZmpC, that selectively induces ectodomain shedding of MUC16, leading to loss of glycocalyx barrier function and enhanced bacterial internalization (11, 12). Under homeostatic conditions, the barrier function of MUC16 in the epithelial glycocalyx is reinforced by galectin-3, a multimerizing lectin that causes carbohydrate-dependent crosslinking of transmembrane mucins (13–15). Intriguingly, galectin-3 is highly expressed in the human cornea and serves as a ligand for P. aeruginosa lipopolysaccharide (16). A potential explanation for the scarcity of adhesive events leading to P. aeruginosa infection in the healthy eye is that the strong, high-affinity association between mucins and galectin-3 on the glycocalyx provides steric hindrance, therefore interfering with the ability of the bacterium to interact with the lectin.

Some bacteria exploit proteoglycans present on the ocular surface epithelia and underlying extracellular matrix to promote infection. Syndecan-1 and perlecan are two proteoglycans containing chains of heparan sulfate and chondroitin sulfate—major glycosaminoglycans found in the human cornea (17). Syndecan-1 is known to enhance the attachment of S. aureus to the plasma membrane of several cell types but fails to do so in corneal epithelial cells. Instead, S. aureus induces shedding of syndecan-1 from the glycocalyx of corneal epithelial cells to produce ectodomains that interfere with the capacity of neutrophils to kill the bacteria (18). Similarly, the ability of P. aeruginosa and S. pneumoniae to infect the cornea is not associated with direct adhesion to proteoglycans present on the epithelial glycocalyx. P. aeruginosa preferably binds to perlecan exposed in the basement membrane after corneal injury (19, 20), whereas S. pneumoniae adhesion relies on the ability of syndecan-1 to promote the assembly of fibronectin fibrils in the basement membrane (21).

The clearance of bacteria can be enhanced by glycans and glycoconjugates present in the tear fluid bathing the ocular surface epithelia. For example, it has been shown that binding of P. aeruginosa to N-glycans on tear glycoproteins functions as a protective mechanism that reduces bacterial adhesion and infection, most likely by facilitating the removal of the bacteria through the tear drainage system (22). Similarly, studies have shown that surfactant protein D, a collagen-containing calcium-dependent lectin with ability to bind lipopolysaccharide from Gram-negative bacteria, is present in the tear fluid and protects against P. aeruginosa invasion (23, 24).

Like bacteria, many viruses causing infectious disease employ glycans on cell surface glycoproteins and glycolipids to access cells. These viruses often recognize unique glycan signatures on the glycocalyx, which frequently leads to specific tissue and species tropisms (25). An example is human adenoviruses (HAdV), one of the most common causes of ocular infection. Dozens of HAdVs, classified into seven species, A to G, have been identified on mucosal surfaces, but only a limited number of them cause epidemic keratoconjunctivitis in the eye (26). It appears that some of these viruses, primarily those belonging to species D, use sialic acid-containing glycans to establish infections at the ocular surface (27). One of them, the cornea-tropic HAdV-D37, specifically binds a branched oligosaccharide present in glycoproteins containing the GD1a glycan motif and featuring two terminal sialic acids (28). Coxsackievirus A24 variant (CVA24v) is another highly contagious infective agent that causes acute hemorrhagic conjunctivitis. Binding and infection experiments using corneal cells indicate that the cell surface receptor used by CVA24v is composed of a sialylated disaccharide (Neu5Acα2,3Gal) on O-glycosylated proteins (29). Consequently, derivatives based on sialic acid have been developed and shown to be effective in preventing HAdV-D37 and CVA24v binding and infection of human corneal epithelial cells (30, 31).

Efforts have been made to establish how these viruses bypass the transmembrane mucin-rich glycocalyx of the ocular surface epithelium to trigger infection and inflammation. The data suggest that, for successful infection, HAdVs need to degrade components of the mucin barrier. This is exemplified by HAdV-D37, which in contrast to HAdV-D19p, a virus that does not cause epidemic keratoconjunctivitis, releases the MUC16 ectodomain from corneal and conjunctival epithelial cells, causing a decrease in glycocalyx barrier function (32). On other occasions, transmembrane mucins contribute to masking viral entry mediators on the epithelial glycocalyx. Affinity assays have shown that herpes simplex virus type 1 (HSV-1), but not HSV-2, binds galectin-3, a component of the human corneal epithelial glycocalyx. Exposure of epithelial cell cultures to transmembrane mucin isolates decreases HSV-1 infectivity, suggesting that the strong association between transmembrane mucins and galectin-3 in the glycocalyx functions to mask the lectin and to provide protection against viral infection (33).

Many pathogens use glycosaminoglycans on the glycocalyx of host cells to initiate infection, and viruses are no exception. Heparan sulfate serves as a main HSV-1 entry receptor in the cornea and facilitates the development of herpetic keratitis (34, 35). Important to infection is the release of the viral progeny from the infected cells so that infection can disseminate into new target cells. Opposing this process are heparan sulfate chains on parent cells, which trap the exiting viral progenies and inhibit their release. Heparanase, a heparan sulfate-degrading enzyme, plays an essential role in facilitating viral egress. Following herpes infection of human corneal epithelial cells, heparanase promotes a continuous loss of heparan sulfate from the cell surface, leading to virus exit and subsequent tissue damage (36). This mechanism involves the upregulation of corneal epithelial heparanase by herpesvirus to promote enzymatic activity at the cell surface, as well as translocation of the enzyme to the nucleus, where it regulates downstream signaling pathways (37). The interaction of HAdV-D37 with sulfated glycosaminoglycans has also been recently reported. In these experiments, removal of heparan sulfate by heparinase III reduced HAdV-D37 binding to corneal epithelial cells but, at the same time, enhanced viral infection, leading the authors to hypothesize that glycosaminoglycans act as decoy receptors (38). Consequently, efforts have been made in the drug development field to use glycosaminoglycan mimetics as artificial decoy receptors that can inhibit HAdV-D37 cell attachment and infection (39).

Fungal infections of the cornea can have devastating consequences if not treated promptly. They are frequently caused by species of Fusarium, Aspergillus, Curvularia, and Candida, with trauma being the most important predisposing cause (40). The first step for a successful infection is the presence of cell wall components and parasitic adhesins that mediate adhesion to host cells and components of the extracellular matrix. A number of interactions with host cells are known to lead to pathogen internalization, but the studies describing this invasion process remain scarce (41). Studies in corneal epithelium have revealed that Cephalosporium curvulum and Aspergillus oryzae use fucose-specific lectins to gain access to the host cell surface and, subsequently, promote infection and disease (42). It is worth noting here that a significant number of terminal and core fucose structures are present in the differentiated corneal epithelial glycocalyx, which could facilitate this type of interaction (43). In addition to immune cells, the corneal epithelium expresses lectins that also interact with polysaccharides on the fungal cell wall. An example is the C-type lectin dectin-1, which recognizes β-glucan. Binding of dectin-1 to Candida albicans initiates protective responses in the epithelium that include the regulation of the innate immune response through the dectin-1/NF-κB signaling pathway (44).

Acanthamoeba keratitis is a rare but serious parasitic infection of the cornea that can cause permanent vision loss. As previously stated for other microbes, adhesion to the surface of the host is one of the first steps in the pathogenesis of infection. Adhesion of Acanthamoeba is followed by lysis of corneal epithelial cells, degradation of extracellular matrix and penetration into the deeper layers of the cornea (45). A major virulence factor of this parasite is a mannose-binding protein that recognizes mannose-containing glycoproteins on the surface of the cornea and plays a role in promoting cytopathic effects (46). The presence of antibodies against the mannose-binding protein in tear fluid is thought to provide protection by inhibiting the adhesion of the parasite. Importantly, oral immunization with recombinant mannose-binding protein in an animal model has been shown to increase the levels of antibodies against this protein in tears and provide protection against Acanthamoeba keratitis (47).