PATHOLOGICAL CHANGES IN KIDNEYS OF INTENSIVELY BRED BROOK TROUT

~ehulka J.: Pathological Changes in Kidneys oj Intensively Bred Brook Trout (Salvelinus Jontinalis Mitchill, 1815). Acta vet. Bmo, 47, 1978: 67-77. Description is given of histopathological changes in kidneys of juvenile and generative brook trout (Salvelinus Jontinalis Mitchill, 1815) under conditions of intensive rearing and feeding with granulated food mixture and meat scraps. Epithelial cells of renal tubuli showed necrosis and dystrophic calcification with subsequent giant cell granulomatous reaction and formation of stones consisting of brushite CaH(PO.) . 2 H 20, as proved by roentgenometric analysis. An unknown detrimental substance is taken into consideration as pathogenetical agent. Salvelinus Jontinalis, visceral granulomatosis, necrotizing nephrosis, nephrocalcinosis, nephrolithiasis, brushite. Connected with intensive breeding of Salmonids, there is a rising number of cases reported as nephrocalcinosis (Besse, Kinkelin and Levaditi 1968), visceral granulomatosis (Dunbar and Herman 1971), and necrotizing nephrosis (Vitovec, Vladik and Cervinka 1974). In Salmo gairdneri suffering from nephrocalcinosis, white deposits of tricalciumphosphate were described. They were oriented towards the walls of tubuli and surrounded by resorptive granulomas. Corynebacteria were isolated from some of the affected kidneys. The authors tend to consider metabolic calcinosis as the ethiological factor. The disease appeared when sea fish was replaced by pelleted food. Visceral granulomatosis in Salvelinus Jontinalis, in addition to typical changes on visceral organs, is characterized by granulomas and tubular degeneration resulting in lithiasis in the caudal part of kidneys. The authors, in agreement with Wood and Yasutake (1956) in Dunbar and Herman (1971), failed to isolate any infective agent. They concluded that the disease is an inflammatory reaction to irritating chemical substances in food. With necrotizing nephrosis in Salvelinus Jontinalis, dystrophical calcification of desquamated epithelia was described together with a secondary granulomatous reaction to foreign bodies. No grampositive immobile diplobacilli were isolated. Sulfonamides were taken into consideration as etiological factor. The present report is limited to description of the culminating pathological process in the kidney of Salvelinus Jontinalis and the resulting grave nephrolithiasis. Materials and Methods Post-mortem examination was performed in one generation (4+) brook trout (Salvelinus Jontinalis Mitchill, 1815) of 440 mm length and 1200 g mass, and one juvenile (1+) brook trout of 180 mm and 75 g, fed with pelleted food mixture of inland provenience and with meat scraps. The kidney tissue was fixed in 10% neutral formol, decalcified in 0.5 M Che1aton III solution at pH 7-8, and, embedded in paraffin wax. Slides were stained by hematoxyline-eosine, the PAS method, Masson's green trichromium, for reticuline by Gomori's, for calcium by Kossa's, for iron by Perls' and for microbes by Gram's method.

granulomas.Corynebacteria were isolated from some of the affected kidneys.The authors tend to consider metabolic calcinosis as the ethiological factor.The disease appeared when sea fish was replaced by pelleted food.
Visceral granulomatosis in Salvelinus Jontinalis, in addition to typical changes on visceral organs, is characterized by granulomas and tubular degeneration resulting in lithiasis in the caudal part of kidneys.The authors, in agreement with Wood and Yasutake (1956) in Dunbar and Herman (1971), failed to isolate any infective agent.They concluded that the disease is an inflammatory reaction to irritating chemical substances in food.
With necrotizing nephrosis in Salvelinus Jontinalis, dystrophical calcification of desquamated epithelia was described together with a secondary granulomatous reaction to foreign bodies.No grampositive immobile diplobacilli were isolated.Sulfonamides were taken into consideration as etiological factor.
The present report is limited to description of the culminating pathological process in the kidney of Salvelinus Jontinalis and the resulting grave nephrolithiasis.

Materials and Methods
Post-mortem examination was performed in one generation (4+) brook trout (Salvelinus Jontinalis Mitchill, 1815) of 440 mm length and 1200 g mass, and one juvenile (1+) brook trout of 180 mm and 75 g, fed with pelleted food mixture of inland provenience and with meat scraps.
The kidney tissue was fixed in 10% neutral formol, decalcified in 0.5 M Che1aton III solution at pH 7-8, and, embedded in paraffin wax.Slides were stained by hematoxyline-eosine, the PAS method, Masson's green trichromium, for reticuline by Gomori's, for calcium by Kossa's, for iron by Perls' and for microbes by Gram's method.
Physico-chemical analyses of concrements were carried out on the diffractometer Chirana GON-3 with a characteristical CoK-alfa radiation, at 28 kV voltage, 14 rnA current intensity, 1 0 per minute goniometer rotations and 600 mm registration shift.
Photographs of the superficial structure of concrements were taken, after coating with gold, by the scanning microscope Jeol, JSM-50 A. *)

Results
The most striking post-mortem finding in the adult fish were protuberancies of a light colour protruding below the renal capsula particularly on the caudal, enlarged part of the kidney (Fig. 1).
Mter cutting the kidney with a grating noise of the knife, a large number of concrements was removed from the kidney tissue.They had the size from sand particle to 5 mm, were of hard consistency, irregular shape and white to faintly yellowish colour (Fig. 2).No alterations were observed in the rest of the organs.
Microscopical examination of the kidney showed enormously dilated tubular spaces, transformed here and there to cystic formations littered with cylindric, mostly desquamated epithelium (Fig. 5).A layer of mesenchymal cells was below the epithelium, and, a fibrous basophilic mass was inside the dilated tubules.In some of them, granulomatous inflammatory reaction with foreign body giant cells could be observed (Fig. 6, 7).
In the juvenile fish, there were different greyish mutually communicating strips on the kidney, resembling marble (Fig. 8).Microscopically, there was an apparent     general reduction of renal tubules.They were of irregular shape, enlarged, deformed and showed a distinctly thickened wall.The tubules were filled with dystrophically altered epithelial cells.In the amassed cells were large deposits of calcium salts.Granulation developed around their crystals and filled partly or totally the tubules.In the granulation tissue, histiocytes rich in eosinophilic plasma predominated.There was a minor admixture of fibroblasts, while foreign body polynuclear cells occurred sporadically (Fig. 9, 10, 12).At intervals, thin diaphragms of renal mesenchyma with evident fibroproductive alterations penetrated and formed partititions inside the renal tubuli (Fig. 11).Apart from the altered tubules, there were also tubules with normal epithelium, filled with granular eosinophilic masses.In some areas of the kidney, interstitium showed fibrous alterations of the haemopoetic tissue where decreased pigmentation could be observed.The eosinophilic mass in the tubules as well as the minute grains in the histiocytes of granulomas were stained by the PAS method.The chemical analysis of concrements was based mainly on the results of diffraction of the roentgenometric examination; Maxima were satisfactorily relevant to tabellar values for brushite (calciumhydrogenphosphate CaH(P04).2 H 2 0, with the exception of the comparatively intensive reflexion of 2.143 d-value which is not listed in Schneider's tables, but is to be found in the atlas (1974).Microscopical examination showed tubular crystals of brushite (monoclinical system) with evidently good fissility (010), reminding of gypsum (Fig. 3, 4).

Discussion
The described morphological alterations in kidneys of both post-mortem examined fish are identical with the changes reported by Vitovec, Vladik and Cervinka (1974) in necrotising nephrosis.The histopathological picture corresponds to findings described with visceral granulomatosis in Salvelinus Jontinalis (Dunbar and Herman 1971) and partly also with nephrocalcinosis in Salrno gairdneri (Besse, Kinkelin and Levaditi, 1968).Similarly as Vitovec et al. (1974), we proved no grampositive diplobacilli in histological sections, while Vladik et al. (1974), succeeded in cultivating them from altered kidneys.
Considering the character of alterations and the failure to find any infective agent (Wood and Yasutake, see Dunbar and Herman 1971), intoxication by some unknown food component seems to be the more probable etiological factor.A detrimental effect of sulphonamides, however, was reliably excluded by scanning performed earlier.

Fig. l .
Fig. l.Enlargement of the caudal part of kidney with numerous concrements in the adult brook trout.

Fig
Fig. 12., Amorphous mass containing calcium surrounded by giant cell reaction and its fibrotic transformation. (g = foreign body giant cell).Kossa, magnification 90 x .