Fasciola gigantica: Ultrastructural cytochemistry of the tegumental surface in newly- excysted metacercariae and in vitro-penetrated juvenile flukes informs a concept of parasite defence at the interface with the host.

Cytochemical staining techniques were carried out en bloc with in vitro excysted and gut-penetrated Fasciola gigantica larvae in order to visualise the glycocalyx of the tegument, a structure which comprises the parasite component of the host-parasite interface, yet is incompletely preserved by conventional fixation and preparation techniques for electron microscopy. Positive reactivity with ruthenium red and periodic acid-thiocarbohydrazine-osmium (PATCO) techniques revealed that the glycocalyx is polyanionic and carbohydrate-rich throughout its depth. It comprises a trilaminate arrangement, with a thin dense zone and fibrillar layer closely apposed to the outer aspect of the apical plasma membrane, invested by an irregular thick mucopolysaccharide capsule. The latter, not recorded in adult flukes, may represent a specific adaptation to facilitate invasion in the face of host immunity, and may also protect the parasite surface from the action of host- and parasite-derived proteases. Early in the invasion of a naïve host, the glycocalyx may be partly responsible for triggering the responses of innate immunity, while later in infection, or when an anamnestic response is initiated in an immunocompetent host, the antibodies and activated lymphocytes of specific acquired immunity are invoked to interact with the parasite surface. The cytochemical properties of the glycocalyx, together with its potential for dynamic turnover due to exocytosis of the T0 tegumental secretory bodies, are likely to aid neutralisation of potentially damaging immune effectors and ensure their removal from the vicinity of the parasite by sloughing in complex with glycocalyx components.

Fasciola gigantica: Comparison of the tegumental ultrastructure in newly excysted metacercariae and in vitro penetrated juvenile flukes indicates intracellular sources of molecules with vaccinal and immunomodulatory potential.

Using in vitro procedures to prepare newly excysted metacercariae and gut-penetrated juvenile Fasciola gigantica, the ultrastructural features of the tegumental syncytium and perikarya of these ephemeral stages in the host-invasion process were compared. The T0-type tegumental cells in newly excysted metacercariae are packed with stored T0 granules which, following transport to the surface membrane of the syncytium, discharge by exocytosis to maintain the glycocalyx. The T0 cells become depleted of T0 granules during the penetration process, shrink in size, and initiate autophagy in the cytoplasm to facilitate metamorphosis from a storage function to active biosynthesis. The novel products appear to include lysosomes which contribute to the autophagosomes, and T1 granules, necessary for maintenance of the glycocalyx and immunoprotection, as the invasion process continues into the host liver. Residual bodies, the end-products of autophagy, are eliminated from the apical membrane of the tegumental syncytium into the host-parasite interface. There they may present a transient source of parasite-derived molecules, including lysosomal cathepsin-type proteases, with potential for interaction with the host's immune system, and so might be exploited as targets for vaccinal and immunomodulatory studies.