Abstract

BackgroundThe category B agent of bioterrorism, Entamoeba histolytica has a two-stage life cycle: an infective cyst stage, and an invasive trophozoite stage. Due to our inability to effectively induce encystation in vitro, our knowledge about the cyst form remains limited. This also hampers our ability to develop cyst-specific diagnostic tools.AimsThree main aims were (i) to identify E. histolytica proteins in cyst samples, (ii) to enrich our knowledge about the cyst stage, and (iii) to identify candidate proteins to develop cyst-specific diagnostic tools.MethodsCysts were purified from the stool of infected individuals using Percoll (gradient) purification. A highly sensitive LC-MS/MS mass spectrometer (Orbitrap) was used to identify cyst proteins.ResultsA total of 417 non-redundant E. histolytica proteins were identified including 195 proteins that were never detected in trophozoite-derived proteomes or expressed sequence tag (EST) datasets, consistent with cyst specificity. Cyst-wall specific glycoproteins Jacob, Jessie and chitinase were positively identified. Antibodies produced against Jacob identified cysts in fecal specimens and have potential utility as a diagnostic reagent. Several protein kinases, small GTPase signaling molecules, DNA repair proteins, epigenetic regulators, and surface associated proteins were also identified. Proteins we identified are likely to be among the most abundant in excreted cysts, and therefore show promise as diagnostic targets.Major ConclusionsThe proteome data generated here are a first for naturally-occurring E. histolytica cysts, and they provide important insights into the infectious cyst form. Additionally, numerous unique candidate proteins were identified which will aid the development of new diagnostic tools for identification of E. histolytica cysts.

Highlights

  • The parasitic protozoan Entamoeba histolytica is the causative agent of amebic colitis and amebic liver abscesses in humans [1,2]

  • A total of 417 non-redundant E. histolytica proteins were identified including 195 proteins that were never detected in trophozoite-derived proteomes or expressed sequence tag (EST) datasets, consistent with cyst specificity

  • We report the identification of 417 non-redundant E. histolytica proteins including 195 proteins that were not identified in existing trophozoite derived proteome or EST datasets, consistent with cyst specificity

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Summary

Introduction

The parasitic protozoan Entamoeba histolytica is the causative agent of amebic colitis and amebic liver abscesses in humans [1,2]. E. histolytica has a simple, two-stage life cycle, consisting of the infective cyst and colon-invasive trophozoite forms. As a result of unknown stimuli in the intestine, trophozoites again can differentiate into cysts (a process known as encystation), which may be excreted in feces to infect other humans. The inability to encyst trophozoites in vitro has severely impaired our knowledge on the infectious stage of E. histolytica. The category B agent of bioterrorism, Entamoeba histolytica has a two-stage life cycle: an infective cyst stage, and an invasive trophozoite stage. Due to our inability to effectively induce encystation in vitro, our knowledge about the cyst form remains limited. This hampers our ability to develop cyst-specific diagnostic tools

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