Norwalk virus-specific binding to oyster digestive tissues

Abstract

The primary pathogens related to shellfish-borne gastroenteritis outbreaks are noroviruses. These viruses show persistence in oysters, which suggests an active mechanism of virus concentration. We investigated whether Norwalk virus or viruslike particles bind specifically to oyster tissues after bioaccumulation or addition to tissue sections. Since noroviruses attach to carbohydrates of the histo-blood group family, tests using immunohistochemical analysis were performed to evaluate specific binding of virus or viruslike particles to oyster tissues through these ligands. Viral particles bind specifically to digestive ducts (midgut, main and secondary ducts, and tubules) by carbohydrate structures with a terminal N-acetylgalactosamine residue in an alpha linkage (same binding site used for recognition of human histo-blood group antigens). These data show that the oyster can selectively concentrate a human pathogen and that conventional depuration will not eliminate noroviruses from oyster tissue.

Figure

Immunohistochemical detection of Norwalk viruslike particles (VLPs) in oyster digestive tissue. A) VLPs in the digestive diverticulum 12 h after seeding sea water with 10⁹ particles. The arrows show immunoreactivity detected in intraepithelial cells. B) VLPs in the digestive diverticulum 12 h after seeding sea water with 10¹² particles. The arrowhead shows immunoreactivity in a phagocyte located in connective tissue, and the arrow shows immunoreactivity in the lumen of a digestive tubule. C) Attachment of recombinant VLPs to secondary ducts of the digestive diverticula after incubation on tissue sections. The arrows show immunoreactivity in the lumen of ducts. D) Attachment of Norwalk virus to a main digestive duct. The arrow shows immunoreactivity in epithelial duct cells. E and F) Attachment (arrows) of recombinant VLPs to digestive ducts without (E) or with (F) periodate treatment of serial tissue sections. G) Binding of VLPs (arrows) to a main digestive duct of VLPs from the H331A mutant capsid protein. H) Lack of binding of the H329A mutant. I) Binding of HPA lectin to the digestive diverticula. Scale bars: A and B, 10 μm; C, E, F, and H, 40 μm; D, G, and I, 20 μm.