HISTOLOGICAL DESCRIPTION OF ENTERIC INFECTION DUE TO THE MICROSPORIDIAN ENTEROSPORA NUCLEOPHILA IN GILTHEAD SEABREAM (SPARUS AURATA)

Emaciative syndrome due to the enteric microsporidian Enterospora nucleophila is an emergent disease of gilthead sea bream (Sparus aurata) farmed in the Mediterranean. During 2015 and 2016 winters, two outbreaks of emaciative syndrome have been observed in caged gilthead sea bream from the Thyrrenian Sea. Sixty-two fish (3.5-110g) were necropsied and subjected to parasitological examination. Gastrointestinal tracts were fixed in 10% buffered formalin and processed for histology. Immunohistochemistry (IHC) was performed on some positive samples using polyclonal antibody anti-Encephalitozoon cuniculi. Intestinal tissue was also subjected to amplification of a fragment of 18S rDNA and sequenced. At necropsy, fish showed emaciation and melanosis; gills and internal organs appeared pale and intestine was often enlarged and filled with yellowish, mucoid exudate. Microscopical examination of intestinal scrapings and wall fresh mounts showed the presence of aggregates of minute spores of microsporidia identified as E. nucleophila by molecular analysis. Histology of stomach and intestine showed in massive infections a strong mucosal sloughing off and necrosis in association to inflammatory infiltration of lymphocytes and few mast cells. In mild infections, intestinal mucosa seemed to be preserved. Microsporidian spores have been detected both in gastric and intestinal mucosa, extending to the lamina propria and the sub-mucosal layer. In advanced phases of infection, the complete destruction of epithelial layers was observed, with thinning of the muscular layer, associated to the presence of conspicuous clusters of infected cells. Spores showed intracytoplasmic or intranuclear localization in enterocytes and only intracytoplasmic localization in macrophages. E. nucleophila spores were strongly positive by IHC. This study confirms the pathogenic role of E. nucleophila in gilthead seabream farmed in the Mediterranean and points out the need of studies aimed to clarify its biology and epidemiology.