Giardia trophozoite-secreted proteins and their effects on intestinal epithelia
Giardia is a major cause of diarrheal disease worldwide. It is a flagellated cyst-forming enteric pathogen that inhabits the lumen of the small intestine. Two genetically distinct lineages (assemblages A and B) are of public health relevance and are often associated with water-borne outbreaks. Yet, the mechanism of pathogenesis and virulence in Giardia is poorly understood. A soluble component derived from healthy, viable and human infective Giardia trophozoites was shown to be able to mediate profound changes in the physiology of human derived enteric cells, consistent with the production of secreted virulence factors by the parasite. Quantitative proteomic analysis was successfully applied to the whole parasite and supernatants derived from the parasite in order to ascertain which parasite proteins are secreted. The genome of Giardia is believed to contain open reading frames which could encode as many as 6,000 proteins although hitherto there was only direct evidence for expression of a few hundred of these. Approximately 1,600 proteins were identified from each assemblage, the vast majority of which being common to both lineages. To look for actual enrichment in the supernatant, the ratio of proteins in the supernatant was compared with the pellet. This defined a far smaller group of putatively secreted proteins enriched comprising a high proportion encoded by genes annotated to have signal peptides, known virulence factors such as the Cathepsin B cysteine proteases and Variable Surface Proteins, scavenging proteins such as an extracellular nuclease and a high proportion of hitherto hypothetical proteins and proteins of unknown function. Further analysis of the genes encoding these proteins indicated that they were highly variable and likely to be under positive selection pressure, confirming their probable role in host-pathogen interactions and their potential as markers for discriminating virulent strains. Based on the proteomic analysis, a new model of pathogenic mechanism for Giardia-induced damage to enteric epithelium in which extracellular nuclease, Cathepsin B and Tenascin may have a concerted action was proposed and may have important implications in the understanding of Giardia pathogenesis.
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