Figure 5. Progressive increase in the size of proteinase K resistant deposits in BSE-challenged fish. The mean diameter of the immunohistochemically detected (SaurPrP1) deposits after protein- ase K–digestion is given with reference to time p.i.. doi:10.1371/journal.pone.0006175.g005 states (Fig. 8C, F, I, L) that cannot be easily classified into any of the previously described morphological categories. The classical neuropathological hallmarks of prion diseases include neurodegeneration, spongiform change and gliosis, while PrPSc deposition is observed in the majority of TSEs. The lesion profile in the brains of the BSE-challenged fish shares both similarities and differences in comparison to this histological and immunohistochemical pattern of mammalian prion diseases. Specifically, the distribution of abnormal aggregates within the brains of 4 BSE-challenged fish at 24 months p.i. shared certain similarities with the PrPSc deposition pattern observed in TSE- TSE Transmission to Sea Bream affected mammalian brain. Notably, the abnormal deposits in sea bream brain were only detected in regions where neuronal parenchyma was present, a feature greatly resembling the location of mammalian prion deposition [36]. Cerebellum was extensively affected, with large fibrous aggregates in the molecular layer and a granule-like deposition profile in the granular layer, a pattern similar to that observed in mammalian TSEs [37]. The abnormal deposition in sea bream brain was also prominent both in the lateral nucleus of the ventral telencephalic area, a fish counterpart to the basal nucleus of Meynert in mammals [38], and the lateral telencephalic pallium, homologue to the mammalian hippocam- pus [22]. Both thalamus and diencephalon displayed numerous aggregates, most of which were interspersed within neuronal fibers. In striking contrast to the general neuropathological profile of mammalian TSEs, however, no vacuoles were observed in any regions of the fish brains examined. While spongiosis is a main characteristic in most prion diseases, it must be noted that in certain TSE subtypes there is little or no spongiform change. Such has been the case in patients suffering from FFI, an inherited human prion disease [39]. Evidence of neurodegeneration, although distinct from that commonly associated with mammalian prion disease, was apparent in many brain regions, primarily in places where the abnormal deposition was located adjacent to or within complexes of neurites (Fig. 8C, L). It is important to note, however, that no degeneration associated with neuronal somata was detected in any of the anatomical regions examined. The absence of classically defined neurodegeneration might be related to the ability of fish to produce new neurons continuously throughout their lifetime. It is known, for instance, that adult fish can regenerate damaged retinal tissue, optic axons and descending brainstem axons, leading to functional recovery [40,41]. In fact, this ability of adult fish for CNS regeneration has been postulated to explain the asymptom- atic carrier state of halibut persistently infected with nodavirus Figure 6. Congo red staining of deposits in the brain of BSE-challenged sea bream. A sagittal, 10 mm-brain section from a BSE-challenged individual, 24 months p.i., was stained with Congo red. A, Diencephalon with light microscopy; B, Same region in polarized light. Rectangles indicate areas of magnification shown in the panel directly below. Arrowheads indicate the abnormal aggregates. Scale bars, 100 mm. doi:10.1371/journal.pone.0006175.g006 PLoS ONE | www.plosone.org 7 July 2009 | Volume 4 | Issue 7 | e6175