BackgroundAlthough the ultrastructure of the schistosome esophageal gland was described >35 years ago, its role in the processing of ingested blood has never been established. The current study was prompted by our identification of MEG-4.1 expression in the gland and the observation of erythrocyte uncoating in the posterior esophagus.Methodology/Principal FindingsThe salient feature of the posterior esophagus, characterized by confocal and electron microscopy, is the enormous increase in membrane surface area provided by the plate-like extensions and basal invaginations of the lining syncytium, with unique crystalloid vesicles releasing their contents between the plates. The feeding process was shown by video microscopy to be divided into two phases, blood first accumulating in the anterior lumen before passing as a bolus to the posterior. There it streamed around a plug of material revealed by confocal microscopy as tethered leucocytes. These were present in far larger numbers than predicted from the volume of the lumen, and in varying states of damage and destruction. Intact erythrocytes were detected in the anterior esophagus but not observed thereafter, implying that their lysis occurred rapidly as they enter the posterior. Two further genes, MEGs 4.2 and 14, were shown to be expressed exclusively in the esophageal gland. Bioinformatics predicted that MEGs 4.1 and 4.2 possessed a common hydrophobic region with a shared motif, while antibodies to SjMEG-4.1 showed it was bound to leucocytes in the esophageal lumen. It was also predicted that MEGs 4.1 and 14 were heavily O-glycosylated and this was confirmed for the former by 2D-electrophoresis and Western blotting.Conclusions/SignificanceThe esophageal gland and its products play a central role in the processing of ingested blood. The binding of host antibodies in the esophageal lumen shows that some constituents are antibody targets and could provide a new source of vaccine candidates.

The surface tegument of the liver fluke Fasciola hepatica is a syncytial cytoplasmic layer bounded externally by a plasma membrane and covered by a glycocalyx, which constitutes the interface between the parasite and its ruminant host. The tegument's interaction with the immune system during the fluke's protracted migration from the gut lumen through the peritoneal cavity and liver parenchyma to the lumen of the bile duct, plays a key role in the fluke's establishment or elimination. However, little is known about proteins of the tegument surface or its secretions. We applied techniques developed for the blood fluke, Schistosoma mansoni, to enrich a tegument surface membrane preparation and analyse its composition by tandem mass spectrometry using new transcript databases for F. hepatica. We increased the membrane and secretory pathway components of the final preparation to ∼30%, whilst eliminating contaminating proteases. We identified a series of proteins or transcripts shared with the schistosome tegument including annexins, a tetraspanin, carbonic anhydrase and an orthologue of a host protein (CD59) that inhibits complement fixation. Unique to F. hepatica, we also found proteins with lectin, cubulin and von Willebrand factor domains plus 10 proteins with leader sequences or transmembrane helices. Many of these surface proteins are potential vaccine candidates. We were hampered in collecting tegument secretions by the propensity of liver flukes, unlike blood flukes, to vomit their gut contents. We analysed both the 'vomitus' and a second supernatant released from haematin-depleted flukes. We identified many proteases, some novel, as well as a second protein with a von Willebrand factor domain. This study demonstrates that components of the tegumental surface of F. hepatica can be defined using proteomic approaches, but also indicates the need to prevent vomiting if tegument secretions are to be characterised.

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