Porencephaly and Periventricular Encephalitis in a 4-month-old Puppy: Detection of Canine Parvovirus Type 2 and Potential Role in Brain Lesions.

A 4-month-old puppy died after showing intracranial signs a few days after a suspected viral enteritis. Grossly, the right cerebral hemisphere had a large irregular cavity external to the internal capsule. Histopathological examination revealed a cystic lesion in the right hemisphere and non-suppurative inflammation of the diencephalon and periaqueductal nervous tissue. Porencephaly associated with periventricular non-suppurative encephalitis was diagnosed. A nested polymerase chain reaction (PCR) identified the presence of parvovirus DNA in the brain and real-time PCR typed this as canine parvovirus (CPV) type 2a. Immunohistochemistry revealed the presence of CPV antigen in the cytoplasm of scattered cells in the subependymal layers and choroid plexus epithelium. The porencephaly was not associated with inflammatory lesions or CPV antigen and was considered to have preceded the neurological signs. In contrast, the detection of CPV antigen in the subependymal layers and choroid plexus epithelium supported the association of this virus with the periventricular encephalitis.

Abortion in goats by Caprine alphaherpesvirus 1 in Spain.

An abortion outbreak occurred in a goat herd of Murciano-Granadina breed in Almeria Region in Spain where 80 pregnant females aborted. All bacteriological and parasitological examinations resulted negative, whereas virological investigations and real-time PCR assay showed the presence of Caprine alphaherpesvirus 1 DNA in the pathological specimens from aborted foetuses. Nucleotide sequence analysis revealed that the DNA was highly close related to the Swiss strain E-CH (99.7%) and a little less extent to the Italian BA.1 strain (99.4%). Histopathological examination revealed multifocal, well-circumscribed, 50- to 200-µm-diameter foci of coagulative necrosis in the liver, lungs and kidneys of three foetuses. In the periphery of the necrosis, there were frequently epithelial cells with the chromatin emarginated by large, round, amphophilic intranuclear viral inclusion bodies. The source of the infection in the herd could not clearly find out even some hypothesis were formulated. This seems to be the first report of an abortion outbreak due to Caprine alphaherpesvirus 1 in a goat herd in Spain.

Histiocytic sarcoma of the nasal cavity in a horse.

Histiocytic diseases in veterinary medicine have been revised in the last few decades, but these are considered relatively rare in horses. This report describes a 9-year-old female horse, Dutch Warmblood, presented for investigation of severe nasal bleeding. A multinodular bilateral mass of 5 cm, reddish to white in color, that invaded and destroyed the surrounding tissues, was observed during a clinical examination of the nostril The morphological features of the tumor cells were represented by cytologically bizarre, highly phagocytic, multinucleated giant cells. These findings, together with immunohistochemical results allowed a diagnosis of histiocytic sarcoma.

Apoptosis of gut-associated lymphoid tissue in rainbow trout Oncorhynchus mykiss after incubation with Candida albicans and bacterial lipopolysaccharide.

Until now a few studies have been carried out on the gut lymphoid system in fish despite its protective role in the host. Here, we have evaluated the effects of Candida albicans (Ca) and lipopolysaccaride (LPS) on the pyloric and terminal segments of gut in the rainbow trout Oncorhynchus mykiss. In particular, data show that both Ca and LPS are able to cause apoptosis of intestinal lymphoid cells as detected by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) procedure. These findings suggest a further modality of gut response in fish to environmental antigens.

In vitro elicitation of intestinal immune responses in teleost fish: evidence for a type IV hypersensitivity reaction in rainbow trout.

In fish the gut immune system has been the subject of few investigations until now. Here, we provide novel morphological and immunological data on the gut isolated from rainbow trout Salmo gairdneri. The pyloric (P) and terminal (T) segments of trout gut, when morphologically examined, evidenced lymphocytes and macrophages (MØ) loosely dispersed in the intestinal mucosa and in the lamina propria in the absence of typical Peyer's patches-like structures. Furthermore, incubation of P and T sections with Candida albicans (Ca) and functional analysis of supernatants generated some interesting results. In fact, active supernatants, when compared with controls, exhibited cytokine-like activities attributable to the presence of interferon (IFN)-gamma and migration inhibiting factor (MIF), respectively. In particular, IFN-gamma-like activity gave rise to an enhancement of Ca phagocytosis by MØ, whereas MIF inhibited MØ migration in agarose. Taken together, these in vitro data suggest that the gut-associated lymphoreticular tissue in fish possesses the appropriate armamentarium to mount a type IV hypersensitivity response when challenged by microbial antigens.

Antigenically activated avian erythrocytes release cytokine-like factors: a conserved phylogenetic function discovered in fish.

Fish erythrocytes are endowed with the ability to produce cytokine like factors when stimulated with Candida albicans (Ca). To evaluate whether similar activities are still conserved in bird erythrocytes (BE), a morphological, cytochemical and immunological evaluation was conducted on peripheral cells in chickens (Gallus gallus). BE form rosettes with monocytes (Mo)-macrophages (MØ), and Mo-MØ according to cytochemical analysis to maintain phagocytic functions across the evolution. Finally, Ca-activated BE release in the supernatants cytokine like-factors that enhance Mo-MØ phagocytosis (interferon-gamma-like activity) and inhibit Mo-MØ migration in agarose (migration inhibitory factor activity). In conclusion, bird erythrocytes, as nonimmune cells, are able to participate in the immune response contributing to the host defence.

First observation of an Hashimoto thyroiditis-like disease in horses from Eastern Europe: histopathological and immunological findings.

Among 622 slaughtered horses from eastern Europe, 156 thyroid glands were selected on the basis of macroscopic alterations (e.g., determination of volume and weight). In the 80% of these thyroids, microscopic alterations consistent with a diagnosis of Hashimoto thyroiditis-like disease were found. In particular, a subverted architecture of the thyroid gland with colloid rarefaction, lymphocytic infiltration, and fibrosis was noted. The confirmation of the histopathological diagnosis of an equine Hashimoto thyroiditis-like disease was provided by the increased serum concentration of thyroglobulin, of antithyroglobulin, and of antithyroid peroxidase autoantibodies. Despite evidence consistent with an Hashimoto thyroiditis-like disease in eastern European horses, the etiopathogenesis of this autoimmune disorder deserves further investigation. In this respect, in some horses histopathological alterations in the pituitary gland may suggest an as-yet-unidentified disorder within the hypothalamus-pituitary adrenal axis associated with Hashimoto thyroiditis.

Modifications of serum and cellular parameters in trotters after a race. Macrophage migration inhibitory activity reduction and serum beta-glucan elevation.

Trotters are exposed to a chronic prolonged stress, such as daily training and frequent races during their active lifespans. There is evidence that trotters undergo very often lethal lung infections after a race, and therefore, is likely that modifications of certain physiologic cellular parameters could account for the increased susceptibility to microbial diseases. Here, we demonstrate that in 7 trotters after a race either serum values (e.g., glycaemia, triglycerides, transaminases, gamma-glutamyltransferase, cholinesterase, amylase, alkaline phosphatase, total proteins, serum albumin, sodium, blood urea nitrogen, lactic dehydrogenase, creatine phosphokinase, and creatinine) or hematological parameters (red blood cell count, hemoglobin, lymphocyte and monocyte count) were increased. At the same time, in the same animals after a race, macrophage migration inhibitory factor activity was depressed, thus indicating an impaired T-lymphocyte response. Finally, increased levels of circulating beta-glucans in some horses, after a race, may suggest a reduced clearance of fungal cell wall components. Taken together, these findings indicate a condition of multiple organ dysfunction, such as the liver, the kidney, the pancreas, and skeletal muscles, as well as a reduced cell-mediated immune response in trotters, after a race.

Do fish thrombocytes play an immunological role? Their cytoenzymatic profiles and function during an accidental piscine candidiasis in aquarium.

Fish (F) thrombocytes (THRs) from healthy trouts were studied in terms of cytoenzyme expression. FTHRs were positive to acid periodic of shiff (PAS) and acid phosphatase (ac. phos.) without tartaric acid (-TA) stainings, as well to alkaline phosphatase. However, when compared with autologous macrophages (M psi's), they were negative to naphthol cloroacetate esterase (AS-D), alpha-naphthyl acetate esterase (Anae), peroxidase (perox) and control ac. phos. with tartaric acid (+TA) stainings, thus indicating a lack of typical lysosomial enzymes. This evidence supports the notion that FTHRs are not true digesting cells. Quite interestingly, trouts and human M psi's were positive for PAS, AS-D, Anae, and perox stainings, thus confirming that cellular cytochemistries are maintained across evolution as their phagocytic functions. Additionally, blood films from trouts, accidentally infected with Candida albicans in aquarium, were morphologically analyzed. Actually, FTHRs interact with erythrocytes, potentiating the formation of rosettes around a central Mpsi. Polymorph nuclear cells and lymphocytes are present in these cellular aggregates, thus suggesting that FTHRs may represent a link between innate and adaptive immunity.

Lymphoreticular system in fish: erythrocyte-mediated immunomodulation of macrophages contributes to the formation of melanomacrophage centers.

In fish, melanomacrophage centers (MMC) represent the equivalent of germinal centers of birds and mammals. They consist of aggregates of macrophages filled with fragments of cells, mainly erythrocytes, and various pigments such as hemosiderin, lipofuscin, and melanin. The kidney, the spleen, and the liver contain many MMC either moving freely or encapsulated in the context of the lymphoreticular tissue. Here, we have demonstrated for the first time that MMC exist in the kidney, spleen, and liver of the rainbow trout Salmo gairdneri Richardson by using traditional stainings and cytoenzymatic methods: periodic acid-shiff, alpha-naphthyl acetate esterase, and peroxidase. Finally, in the view of the already described peripheral interaction between fish erythrocytes (FE) and macrophages (MO), we postulate that these cellular aggregates (rosettes) travel to distant organs, thus contributing to the formation of MMC in the kidney, spleen, and liver.

Maturation of fish erythrocytes coincides with changes in their morphology, enhanced ability to interact with Candida albicans and release of cytokine-like factors active upon autologous macrophages.

Erythrocytes from the rainbow trout Salmo gairdneri Richardson (Salmo g.R.) were classified into immature and mature populations, respectively, by measuring longitudinal diameters. More elongated fish erythrocytes (FE), classified as mature cells, were those interacting with Candida albicans (CA) in a higher frequency in terms of either binding to the fungus or its intracellular engulfment. At the same time, in the rosetting phenomenon more elongated mature FE surrounded macrophages (Mø) phagocytosing CA. Finally, FE activated by CA released in the supernatants cytokine-like factors able to modulate Mø functions. In particular, these active supernatants were analyzed for their capacity to inhibit Mø migration Macrophage Inhibition Factor (MIF) activity and enhance Mø phagocytosis. Both activities were detected in supernatants from CA stimulated FE but not in control supernatants. MIF activity could play a role in the accumulation of Mø in the context of functional rosettes, while the factor enhancing Mø phagocytosis could promote clearance of CA in a more efficacious way.

A phylogenetic comparison between acute monocytic leukemia cells and monocytes-macrophages in lower vertebrates.

In humans, monocytes and macrophages (Mphi) play a central role in immune regulation, tissue maintenance and pathogen control. In lower vertebrates, a few studies have been conducted on Mphi like cells. In acute monocytic leukemia monocytic cells, as immature cells restrained in one of the phases of their ontogenesis, would offer the opportunity to rebuild an archaic condition helpful to understand the phylogenesis. Therefore, aim of this work was to characterize in the Rainbow trout (Salmo Gairdneri Richardson) Mphi and compare them with acute leukemia monocytic cells. In the trout, Mphi's morphology is similar to that of mammals. In particular, Mphi possess an irregular embryoshaped nucleus occupying 2/3 of the cell, while the peripheral cytoplasmic profile is irregular with extroflexed plasmalemma and pseudopods. A morphological transition towards Mphi is featured by a wavy hyaline classical membrane and an irregular and extroflexed surface. Some aspects of erythrophagocytosis represented a finding of great interest indicating that the hemocatheretic function could take place directly in circulation. This condition, also observed in human acute monocytic leukemia, suggests that the information to the erythrophagocytosis is restrained under physiological conditions. Non-specific esterases, which are positive in human Mphi smear and Mphi from human lymph node tissue, were also positive in the teleost studied but with a dysomogeneous pattern. Consequently non-specific esterase system is phylogenetically conserved. A lack of immune-reactivity with the anti-CD68 monoclonal antibody (MoAb) on smear and trout tissue sections was observed. On the contrary, strong positivity was detected on human lymph node sections. In trout, the presence of Mphi and circulating Mphi like cells exhibiting an erythrocatheretic function in the circulation would indicate a primordial function that has later been replaced by the liver and the spleen.

Fish immunology. II. Morphologycal and cytochemical characterization and phagocytic activities of head kidney macrophages from rainbow trout (Salmo gairdneri Richardson).

In Salmo gairdneri Richardson trouts, a comparison was made between macrophages (MO) derived from head kidney and peripheral blood monocytes. Morphologically and cytochemically no differences were observed between these two types of mononucleated cells. On the other hand, in parallel studies the ability of trout erythrocytes to engulf Candida albicans (CA) was evaluated and compared to the MO phagocytosis. In erythrocytes, engulfment is preceded by binding to CA and cell membrane invagination, while this was not the case for MO. Finally, MO stimulated with lipopolysaccharides (LPS) did not modify their phagocytic capacities, thus suggesting a lack of LPS receptors or a tolerance to LPS.

Fish immunology. I. Binding and engulfment of Candida albicans by erythrocytes of rainbow trout (Salmo gairdneri Richardson).

The role of fish erythrocytes (FE) as phagocytic cells has poorly been investigated, until now. Here, we have focussed our attention on the interplay between rainbow trout (Salmo gairdneri Richardson) erythrocytes and Candida albicans (CA). At the same time, the intervention of autologous head kidney macrophages (MO) in the CA processing by FE has been studied. Data show that CA particles bind to FE, which, in turn, are able to engulf but not kill them. In the presence of MO, a decrease of FE with bound CA occurs and, in some microscopic images, FE form rosettes with MO. Phagocytosis of CA is higher in rosetting MO than in non-rosetting ones. According to our findings, it appears that FE represent a reservoir of engulfed CA and rosetting is an efficacious phenomenon of presentation of pathogens to MO, where an effective clearance of them can take place.

The rainbow trout (Salmo gairdneri Richardson) granular leukocytes and mast-cells system versus the human one. Cytomorphometrical and cytochemical characters in these two species philogenetically poles apart.

In man, circulating granular leukocytes constitute a cellular system and are able to migrate in the tissues to take part in the immune reactions. This study was to characterize the granular leukocytes and the eventual existence of mast-cells of a low vertebrate, rainbow trout (Salmo gairdneri R.) in order to compare it with man. The blood smears and tissues sections have been tested whit panoptic methods, cytochemistry and toluidine blue. White blood cell count, leukocytes formula and cytomorphometric characterization was performed using an image analyser. Scanning of tissues sections in order to identify mast-cells has been also performed. In this model the granular leukocytes are all neutrophilic like; no eosinophilic, no basophilic no tissue mast-cells, basically existing in allergic and anaphylactic reactions, were found.

Corrosion cast of the vascularization of Mugil Cephalus gills.

The spatial organization of the microcirculation in gills of Mugil Cephalus, was examined by scanning electron microscopic analysis of corrosion cast prepared by intravascular injection of methyl methacrylate. The afferent branchial artery originates from the ventral aorta and gives rise to afferent filamental artery. From the medio-lateral wall of the afferent filamental artery, afferent lamellar arterioles originate which supply one or more lamellae. The lamellar efferent arterioles, which drain the blood coming from the lamellae into the efferent filamental arteries, continue with the efferent branchial arteries and then the dorsal aorta. The techniques used so permitted to evaluate the structure and the interrelationships of the vascular pathways, explaining the regulation and the distribution of the blood flow in the gills.

GABA concentrations in different cerebral areas of bass (dicentrarchus labrax).

In these investigations GABA concentrations in different cerebral areas (cortex, diencephalon, optic lobes and mesencephalon) of bass (Dicentrarchus labrax) were evaluated. According to the results obtained, the GABA, like other chemical mediators (noradrenaline, dopamine, serotonin), investigated in different animal species, shows a variable distribution in the CNS of bass. The highest levels of GABA were observed in the cerebral cortex and the lowest ones in the mesencephalon. Mean values were observed in the diencephalon and the optic lobes. The study of the GABA-ergic system of the CNS of fish, could provide a valid contribution to the comprehension of several biochemical and physiopathological mechanisms responsible for the behavioural activity and/or the appearance of neurotoxic phenomena.

[Comparative anatomo-microscopic observations on the esophagus of teleosts with different feeding habits]. / Osservazioni anatomo-microscopiche comparate sull'esofago di teleostei a diverso regime alimentare.

The authors describe the micro-anatomy of the oesophagus of different diet fish: herbivorous (Boops salpa), omnivorous (Liza aurata) and carnivorous fish (Blennius gattorugine and Gaidropsarus mediterraneus). In all the observed species the oesophagus is formed by a tunica mucosa, tunica muscularis and tunica serosa. The tunica mucosa of the herbivorous fish arises to form thin plicae which protrude into the lumen and are covered with squamous stratified epithelium. The goblet cells do not arrange among the covering cells but take place laterally and on the basis of the plicae. The tunica muscularis is constituted by an external circular layer and an inner longitudinal layer of striated muscle. The oesophagus of omnivorous fish shows thick plicae covered with a higher epithelium than that of the herbivorous fish. Among the epithelial cells a lot of taste buds have been found. The tunica muscularis is like that of the herbivorous fish. In the carnivorous fish Blennius the tunica mucosa presents thin plicae like those of the herbivorous fish but they are covered with a thicker epithelium. The tunica muscularis is formed by an external circular layer and by an inner longitudinal layer which constitutes two diametrically opposed bundles placed dorsally and ventrally. In the other carnivorous fish Gaidropsarus, the tunica mucosa shows very thick plicae covered with a quite high squamous epithelium among which there are many goblet cells. In this fish too numerous taste buds have been found. The tunica muscularis is arranged in the same manner observed in the other carnivorous fish.

[Comparative macroscopic and microscopic anatomical observations on pyloric ceca in some bony fish]. / Osservazioni anatomo-macro-microscopiche comparate sui ciechi pilorici di alcuni pesci ossei.

The authors report the variable number of pyloric caeca of some species of fish and describe their macroanatomy. They also describe and compare the microanatomy of pyloric caeca in three species; in particular they point out the thickness of tunica muscularis, the shape and length of villi which protrude into the lumen and the variable number of goblet cells. The tunica muscularis is very strong in Uranoscopus and consists of three strata: external and inner longitudinal strata, middle circular stratum. The tunica muscularis is less thick in Diplodus and in Scorpaena. The lamina propria and epithelium form villi which protrude into the lumen. They are covered with columnar epithelium and goblet cells which are more numerous in Uranoscopus and Scorpaena and less numerous in Diplodus. At last the authors describe in pyloric caeca of Scorpaena the presence of a network which engages all the lumen; it consists of connective tissue septa which directly derive from the lamina propria and is covered with the same columnar epithelium and goblet cells.

[Further observations and considerations on the biological and reproductive cycle of the roe buck (Capreolus capreolus L.)]. / Ulteriori osservazioni e considerazioni sul ciclo biologico e riproduttivo del capriolo (Capreolus capreolus L.).

The authors have studied the seasonal microanatomical modifications of the ovary of the roe deer and testis of the roe buck. The ovary during the month of September presents primary, secondary and mature vesicular follicles. During the month of October there is the first presence of a corpus luteum. The corpora lutea during the months of November and December increase in size and blood supply; the cells are filled with acidophil granules. The corpora lutea persist also in January and February when embryos are implanted in the uterus. This result agrees with Short's and -Hay's but differs from that of Stieve. Testes show the first spermatogenic activity during the month of April. The spermatogenesis is completed in June, persists and reaches the top during July and August, when the Leydig cells acquire a strong acidophil cytoplasm. Indeed the spermatogenesis decreases during September and October; then, during winter months the testes are devoid of all signs of spermatogenic activity, the tubules being lined only be Sertoli cells and spermatogonia.