Apical Secretory Glycoprotein Complex Contributes to Cell Attachment and Entry by Cryptosporidium parvum.

Cryptosporidium parvum is an enteric pathogen that invades epithelial cells in the intestine, where it resides at the apical surface in a unique epicellular location. Compared with those of related apicomplexan parasites, the processes of host cell attachment and invasion by C. parvum are poorly understood. The streamlined C. parvum genome contains numerous mucin-like glycoproteins, several of which have previously been shown to mediate cell attachment, although the majority are unstudied. Here, we identified the antigens recognized by monoclonal antibody (MAb) 1A5, which stains the apical end of sporozoites and mature merozoites. Immunoprecipitation with MAb 1A5 followed by mass spectrometry identified a heterodimer comprised of paralogous proteins which are related to additional orthologs in the genome of C. parvum and related species. Paralogous glycoproteins recognized by MAb 1A5 heterodimerize as a complex displayed on the parasite surface, and they also interact with lectins that suggest that they contain mucin-like, O-linked oligosaccharides. Although the gene encoding one of the paralogs was readily disrupted by CRISPR/Cas9 gene editing, its partner, which contains a mucin-like domain related to GP900, was refractory to deletion. Combined with the ability of MAb 1A5 to partially neutralize host cell attachment by sporozoites, these findings define a new family of secretory glycoproteins that participate in cell invasion by Cryptosporidium spp. IMPORTANCE Although Cryptosporidium is extremely efficient at penetrating mucus and invading epithelial cells in the intestine, the mechanism of cell attachment is poorly understood. To expand our understanding of this process, we characterized the antigens recognized by a monoclonal antibody that stains the apical end of invasive stages called sporozoites and merozoites. Our studies identify a family of glycoproteins that form heterodimers on the parasite cell surface to facilitate host cell attachment and entry. By further defining the role of mucin-like glycoproteins in host cell attachment, our studies may lead to strategies to disrupt cell adhesion and thereby decrease infection.

Neonatal Mouse Gut Metabolites Influence Cryptosporidium parvum Infection in Intestinal Epithelial Cells.

The protozoan parasite Cryptosporidium sp. is a leading cause of diarrheal disease in those with compromised or underdeveloped immune systems, particularly infants and toddlers in resource-poor localities. As an enteric pathogen, Cryptosporidium sp. invades the apical surface of intestinal epithelial cells, where it resides in close proximity to metabolites in the intestinal lumen. However, the effect of gut metabolites on susceptibility to Cryptosporidium infection remains largely unstudied. Here, we first identified which gut metabolites are prevalent in neonatal mice when they are most susceptible to Cryptosporidium parvum infection and then tested the isolated effects of these metabolites on C. parvum invasion and growth in intestinal epithelial cells. Our findings demonstrate that medium or long-chain saturated fatty acids inhibit C. parvum growth, perhaps by negatively affecting the streamlined metabolism in C. parvum, which is unable to synthesize fatty acids. Conversely, long-chain unsaturated fatty acids enhanced C. parvum invasion, possibly by modulating membrane fluidity. Hence, gut metabolites, either from diet or produced by the microbiota, influence C. parvum growth in vitro and may also contribute to the early susceptibility to cryptosporidiosis seen in young animals.IMPORTANCECryptosporidium sp. occupies a unique intracellular niche that exposes the parasite to both host cell contents and the intestinal lumen, including metabolites from the diet and produced by the microbiota. Both dietary and microbial products change over the course of early development and could contribute to the changes seen in susceptibility to cryptosporidiosis in humans and mice. Consistent with this model, we show that the immature gut metabolome influenced the growth of Cryptosporidium parvumin vitro Interestingly, metabolites that significantly altered parasite growth were fatty acids, a class of molecules that Cryptosporidium sp. is unable to synthesize de novo The enhancing effects of polyunsaturated fatty acids and the inhibitory effects of saturated fatty acids presented in this study may provide a framework for future studies into this enteric parasite's interactions with exogenous fatty acids during the initial stages of infection.