Red mark syndrome of trout (Oncorhynchus mykiss; Walbaum, 1792): Histopathological scoring and correlation with gross lesions.

Red mark syndrome (RMS) is a skin disorder affecting rainbow trout (Oncorhynchus mykiss). The present work aimed to correlate the gross skin lesions affecting 46 fish sampled from farms surveyed for RMS with their microscopic features, identifying histological parameters that may be suggestive of disease progression. Skin lesions were grossly included in one of three categories (types I, II and III) according to the progressive degree of severity. Histological parameters and anti-proliferating cell nuclear antigen (PCNA) tissue immunoreactivity were semi-quantitatively assessed. In the dermis, PCNA-positive lymphocytes, fibroblasts and endothelial cells were indicative of active phlogosis. A significant increase in PCNA-immunoreactive lymphocytes, from gross type I to type III cases, was found only in the hypodermis. The histological parameters significantly associated with the gross lesion severity were progressive loss of the epithelium and scales, recruitment of inflammatory cells in the stratum compactum, loss of architecture of the stratum compactum, perivascular and perineural granulomatous inflammation and increase in lymphocyte infiltration of the muscular layer. In the type II and type III categories, inflammation in the hypodermis and muscle displayed a granulomatous pattern, reinforcing the hypothesis of an immunopathological mechanism. The morphological diagnosis of "deep chronic dermatitis associated to panniculitis and myositis, characterised by lympho-histiocytic and granulomatous reaction" is suggested.

Ultrastructural and biomolecular detection of Rickettsiales-like organisms in tissues of rainbow trout with Red Mark Syndrome.

Red mark syndrome (RMS) and US strawberry disease (US SD) are skin disorders affecting rainbow trout farmed in Europe and USA. The disease etiology has not yet been established. In spite of specific investigations, identifying Rickettsia-like organism (RLO)- and Midichloria-like organism (MLO)-related DNA in affected individuals, these pathogens have never been observed. We performed histological, ultrastructural and biomolecular analysis on skin and spleen samples of trout with RMS. Examination by TEM revealed the presence of intracytoplasmic microorganisms resembling Rickettsiales within macrophages, fibroblasts and erythrocytes. The microorganisms were oval or short rod shaped (400-800 nm in length and 100-200 nm in width) and often showed a cell wall similar to Gram-negative bacteria. PCR analysis for Rickettsiales supported these findings: 53% of affected trout were positive by both PCR and TEM The primers RiFCfw-RiFCrev were used to anneal both the RLO 16S DNA sequence and the MLO 16S DNA sequence. For this reason, and in agreement with previous studies confirming the presence of Rickettsiales-related DNA in trout with RMS, we assume that TEM detected microorganisms morphologically consistent with bacteria belonging to Rickettsiales order and could be considered as possible causative agents of RMS.

Experimental Pseudomonas anguilliseptica infection in turbot Psetta maxima (L.): a histopathological and immunohistochemical study.

Experimental infection with Pseudomonas anguilliseptica was performed both by intraperitoneal (i.p.) and bath route on juvenile turbot (Psetta maxima) in order to evaluate the pathology induced. Turbot was found to be sensitive to i.p. challenge (1.7x10(6) CFU/fish) but no to bath exposure. The i.p. challenge induced septicaemic infection and mortality. Externally, moribund fish showed distended abdomen and pale areas at day 9. The gross pathological internal signs present were abundant ascitic fluid in the peritoneal cavity, pale and enlarged spleen, pale and friable liver, and congestive and dilated gut with yellowish exudates. On histopathological examination, bacterial invasion was common in all the tissues studied but the most prominent pathological changes were observed in gut, spleen and kidney after 7 day with features of necrosis. The immunohistochemical findings support the widespread localization of the bacteria after the i.p. injection since the P. anguilliseptica was detected in spleen from day 1 post injection, in liver, kidney and gut from day 4, in muscle from day 7 and in brain from day 9. The difficulties in infecting healthy fish by bath challenge can be explained by the opportunistic nature of this pathogen.

Immunohistochemical and lectin histochemical analysis of the alpaca efferent ducts.

An immunohistochemical and lectin histochemical study of the efferent ducts was performed in the alpaca. Two types of epithelium, consisting of principal and ciliated cells, were detected on the basis of the different cytokeratins expression and lectin binding pattern. AE1/AE3 and 13 cytokeratin antibodies intensely immunostained the entire cytoplasm of type I PCs, whereas AE1/AE3, but not anti cytokeratin 13, immunoreacted in type II principal cells along the apical, lateral and basal plasma-membrane. The histochemical characterization of the epithelial cells was carried out using a battery of different lectins: Con-A, UEA-I, LTA, WGA, GSA-II, GSA-IB4, SBA, PNA, ECA, DBA, MAL-II and SNA. Sialidase digestion and deglycosilation pre-treatments were also employed. In type I principal cells, the staining of the Golgi zone was interpreted giving evidence for the synthesis and secretion of O- and N-linked oligosaccharides. In particular, alpha-Man/alpha-Glc, GlcNAc, beta-Gal-(1-4)-GlcNAc, Neu5Acalpha2,3Gal and Neu5Acalpha2,6Gal/GalNAc residues were included in both O- and N-linked glycans, whereas alpha-Fuc, beta-GalNAc and alpha-Gal were only found in O-linked oligosaccharides; alpha-GalNac and beta-Gal-(1-3)-D-GalNAc were found subterminal to sialic acid moieties and they were included in O- and N-glycans. In type II principal cells, the lectin staining was observed in the apical cytoplasmic granules and in vacuoles that were interpreted as components of an elaborate endocytotic apparatus specialized for the uptake of particulate material and fluid from the lumen. These results suggest the existence of two structurally different epithelial segments along the ductuli efferentes of the alpaca, with a high degree of compartmentalization of the secretory and absorptive activities.

Differentiation of glycans in equine guttural pouches.

The aim of this study was to investigate the carbohydrate composition of mucosubstances in the equine guttural pouches using conventional histochemical tests in conjunction with glycolytic digestion to degrade different classes of glycosoaminoglycans. In the goblet cells, the mucopolysaccharides contained chondroitin sulfate B, heparin, heparan sulfate and sialic acid residues. The acinar cells also expressed these substances (except for heparin), whereas the ductal cells produced chondroitin sulfate B and sialic acid. Neutral sugars were also found in each cell type. The diversity of the glycocomponents found in the auditory tube suggests that they may have important functional roles. Indeed, the glycosoaminoglycans provide a hydrophilic environment that prevents dehydration and desiccation of the guttural membranes during air passage. Additionally, these glycomolecules may be involved in the pathogenesis of some bacterial disease in horses, such as equine strangles.

Experimental Pseudomonas anguilliseptica infection in turbot Psetta maxima (L.): a histopathological and immunohistochemical study.

Experimental infection with Pseudomonas anguilliseptica was performed both by intraperitoneal (i.p.) and bath route on juvenile turbot (Psetta maxima) in order to evaluate the pathology induced. Turbot was found to be sensitive to i.p. challenge (1.7×106 CFU/fish) but no to bath exposure. The i.p. challenge induced septicaemic infection and mortality. Externally, moribund fish showed distended abdomen and pale areas at day 9. The gross pathological internal signs present were abundant ascitic fluid in the peritoneal cavity, pale and enlarged spleen, pale and friable liver, and congestive and dilated gut with yellowish exudates. On histopathological examination, bacterial invasion was common in all the tissues studied but the most prominent pathological changes were observed in gut, spleen and kidney after 7 day with features of necrosis. The immunohistochemical findings support the widespread localization of the bacteria after the i.p. injection since the P. anguilliseptica was detected in spleen from day 1 post injection, in liver, kidney and gut from day 4, in muscle from day 7 and in brain from day 9. The difficulties in infecting healthy fish by bath challenge can be explained by the opportunistic nature of this pathogen.

Tracheobronchial neuroendocrine carcinoma in a cat.

A tracheobronchial neuroendocrine carcinoma in a 10-year-old cat with a history of coughing, dyspnoea and anorexia is described. Endoscopic examination revealed a yellowish mass protruding into the trachea and partly obstructing the right bronchial lumen. Histological examination of biopsy samples revealed a hypercellular tumour consisting of sheets or ribbons of small hyperchromatic cells, with oval to spindle-shaped nuclei and minimal cytoplasm; nucleoli and mitotic activity were absent. The findings were confirmed on post-mortem examination. The small neoplastic cells were immunolabelled to varying degrees by antibodies against neuron-specific enolase, S-100 protein, synaptophysin and vimentin. Ultrastructural studies revealed scattered neurosecretory granules and scanty cellular junctional complexes, including desmosomes and tonofilaments, in neoplastic cells.

Granular cell tumour affecting the left vocal cord in a dog.

Granular cell tumours (GCTs) have been described in human vocal cords, but not in those of the dog. A single polyp in the left vocal cord, associated with laryngeal oedema and monolateral hyperplasia of the tonsil, was observed endoscopically in a 6-year-old male dog. Cytologically, the mass was characterized by well-defined polygonal cells, with granular cytoplasm and an eccentric nucleus. Histologically, the mass showed sheets of globoid to polygonal cells, filled with numerous diastase-resistant, periodic acid-Schiff-positive granules, and scanty stroma. Ultrastructurally, the granules were represented by heterogeneous lysosomes of variable size, suggesting a diagnosis of GCT. As reported in man, hyperplasia of the laryngeal and tonsillar epithelium and the presence of foci of pearl-like epithelial cells, mimicking invasive squamous cell carcinoma, were observed. Immunohistochemical examination revealed labelling of neoplastic cells for S-100 protein, CD68, collagen IV, and, focally, for glial fibrillary acidic protein, neuron-specific enolase and vimentin. These findings suggested a Schwann cell origin of the GCT.