Proliferative kidney disease in brown trout, Salmo trutta L., and Atlantic salmon, Salmo salar L., parr: histopathological and epidemiological observations

Abstract Proliferative kidney disease (PKD) has been described in cultivated rainbow trout but major outbreaks have not been recognized in other fish species. This paper describes light and electron microscope studies and presents epidemiological data of a disease condition in first summer Salmo salar parr and S. trutta , which is associated with an organism apparently identical to the aetiological agent of PKD. The pathology was found to be similar to that described for PKD in rainbow trout. The epidemiology in the brown trout suggests that exposure after May does not result in infection. The data from the outbreaks in two salmon hatcheries appear complex and suggest different susceptibilities to PKD among salmon originating from different sources.     

赤点石斑鱼病毒性神经坏死症的组织病理和电镜观察

用逆转录-聚合酶链式反应（RT-PCR）检测患病赤点石斑鱼苗，呈Beta诺达病毒阳性。光学显微镜下观察到病鱼的脑、视网膜、脊髓有空泡．在脑部，空泡主要分布在端脑、间脑和小脑。受感染的细胞明显收缩、致密变化和嗜碱性。包涵体常为圆形，大小不一。透射电镜下，在感染细胞的细胞质可观察到含有病毒粒子的致密体。病毒粒子呈等面体，无外膜，直径为25～28nm，随机分布在细胞质或在致密体内排列成品格状。致密体大小不一。偶尔观察到较大致密体的外膜已破裂，病毒粒子被释放到细胞质。     

Proliferative kidney disease: cellular aspects of the rainbow trout, Oncorhynchus mykiss (Walbaum), response to parasitic infection

Proliferative kidney disease (PKD) is an immunopathological condition of salmonid fish, caused by the hyperplastic response of their principal lymphoid tissues to infection with the spores of Tetracapsula bryosalmonae , a myxozoan parasite formerly designated proliferative kidney organism – unknown (PKX). In order to investigate the nature of cells involved in this host response and possible alterations of their functions during parasitic infection the course of PKD was studied by flow cytometry (FCM) techniques, using blood, pronephros and spleen leucocyte populations from rainbow trout infected by intraperitoneal (i.p.) injection with parasitic cells from infected donor fish. The parameters of the cellullar response studied were: cytogram of cell population, lymphoproliferation, phagocytosis, oxidative burst, and non-specific cytotoxicity. The modifications of cell population distribution and function in the PKD-infected fish mainly affected the pronephros cell populations and were coincident with the clinical phase of disease. During this phase, the lymphocytes constituted the major leucocyte cell population and underwent proliferation and were thus responsible for the renal tissue hyperplasia. Meanwhile, phagocytosis and oxidative burst were depressed. These data are in agreement with the patho-epidemiological background of PKD where the enhancement of the fish sensitivity to bacterial infections reflects the impairment of certain cellular defence mechanisms of innate immunity.     

三疣梭子蟹光感受器超微结构在昼夜光周期中的变化

应用透射电镜技术显示，三疣梭子蟹（Portunus tritubereulatus）光感受器由11个小网膜细胞组成。其中，4个小网膜细胞组成了感光部分的远端，另外的7个小网膜细胞组成了感光部分的近端，并成为感光系统的主体。在黎明、正午、黄昏和夜晚4个时间点观察到，其小网膜细胞的结构随昼夜光周期发生节律性的变化。特别以感光部位-感杆束的面积和结构的变化最为明显。另外，在小网膜细胞中的板膜体、膜下储泡囊、色素颗粒等细胞器的形态和数量也随光周期呈现出规律性的变化。结果表明三疣梭子蟹光感受器的结构变化与光照有关。     

Ultrastructural aspects of a serious disease of hatchery reared larval oysters, Crassostrea gigas Thtinberg

Abstract. Ultrastructural observations on a possible aetiological agent and tissue changes in a previously described disease of hatchery reared larval oysters Crassostrea gigas Thünberg are reported. Spherical bodies, implicated in gastro-intestinal impaction, show a strong resemblance to the marine fungus, Hyalochlorella marina . Similarities include a thick outer wall structure, plate-like mitochondria and dense cytoplasm with small vacuoles.
The dense fascicular masses, previously observed histologically at epithelial surfaces, consist of accumulations of irregular electron opaque bodies. Large accumulations of these bodies appear to exacerbate tissue damage and prevent normal velar function. In comparison with the reported virus-like particles in this disease, these dense inclusions and associated lesions are more frequently observed and thus seem to be more important in the disease process.
Notes on normal epithelial surfaces of larval Crassostrea spp. are included.     

Distribution and prevalence of T. bryosalmonae in Austria: A first survey of trout from rivers with a shrinking population

The first evidence of proliferative kidney disease (PKD) in an Austrian river (the River Kamp) was documented in 2016, and no information on the PKD infection status of trout in other rivers was available. Since then, brown trout (Salmo trutta fario) and rainbow trout (Oncorhynchus mykiss) have been collected from rivers in Upper and Lower Austria for different diagnostic purposes. In this study, we summarize the recent findings of a first survey concerning the distribution of Tetracapsuloides bryosalmonae, the causative agent of proliferative kidney disease (PKD), from these samples. Between September 2015 and October 2017, a total of 280 brown trout and 39 rainbow trout were collected from 21 rivers in the provinces of Upper and Lower Austria. T. bryosalmonae was detected by PCR of kidney tissue in 17 of 21 sampled rivers and in 138 of 280 brown trout as well as in 11 of 39 rainbow trout. Pathological signs of PKD (e.g., hypertrophy of the kidney) were observed in 33 analysed brown trout and six rainbow trout samples. No correlations between fish infected by T. bryosalmonae and the parameters size and age class, condition factor, geological origin of the streams and distribution within the river course were found, while positively tested fish are significantly increased at sampling sites exceeding water temperatures of 15°C for median periods of 115 days. The prevalence within the affected streams or stream sections is highly variable, and in single rivers, infection rates of up to 90% are confirmed.     

Light and electron microscope observations on pathological changes in the gills of the marine fish Lycodes lavalaei Vladykov and Tremblay associated with the proliferation of an unidentified cell

Abstract. Pale and swollen gills were observed in 23 of 240 Laval's eelpouts, Lycodes lavalaei Vladykov and Tremblay (Perciformes), taken from several locations off the coasts of Newfoundland and southern Labrador. Light microscopic study of histological sections revealed clusters of unidentified cells ('X-cells') in the basal region of secondary gill lamellae and extending along the latter, between the epithelium and lamellar capillaries. In the electron microscope two types of X-cells were enclosed in a network of cytoplasmic processes from dense 'envelope' cells. Both types of X-cells were bound by a densely-coated limiting membrane and had nuclei with large, dense nucleoli, numerous mitochondria with short vesicular cristae, lipid inclusions, abundant smooth endoplasmic reticulum and small dense inclusions. The Type II X-cells were larger, contained a much greater concentration of lipid and small dense inclusions, and an additional inclusion with a dense core surrounded by a light halo. The X-cells are compared with strikingly similar cells described from epidermal and pseudobranch tumours of marine fish. Although transformation of a host cell by virus or possibly pollution has been suggested, neither the nature nor the origin of the X-cells can be determined.     

Proliferative kidney disease (PKD) in rainbow trout Salmo gairdneri: Further observations on the effects of water temperature

The seasonality of proliferative kidney disease (PKD) and the effects of water temperature on the development of the disease in naturally-infected fingerling rainbow trout, Salmo gairdneri, were investigated. Fingerling rainbow trout became infected from May to October, but those infected in October did not develop clinical disease. In naturally-infected fish subsequently held under laboratory conditions, clinical PKD occurred at 12–18°C, but not at 9°C. The disease progressed more rapidly and was more severe as temperatures increased. On re-exposure to infection, rainbow trout previously held at 9°C developed clinical PKD, whereas in those previously held at 12°C it did not develop. Possible implications of these findings for the control of PKD are discussed.     

Development of proliferative kidney disease in rainbow trout, Oncorhynchus mykiss (Walbaum), following short-term exposure to Tetracapsula bryosalmonae infected bryozoans

The initial site of infection in the fish host for Tetracapsula bryosalmonae , causative agent of proliferative kidney disease (PKD) is poorly understood. Following the recent recognition that freshwater bryozoans harbour the infective stages to salmonid fish, experimental transmission studies were undertaken to investigate (1) the route of entry of the parasite into the fish host and (2) the minimum exposure time required to induce clinical signs of PKD. In-situ hybridization (ISH) studies were carried out on naïve rainbow trout exposed to the naturally infected bryozoan Fredericella sultana for up to 90 min. The sporoplasm of T. bryosalmonae was detected entering the fish via mucous cells in the skin epithelium within the first minute of exposure. In addition, T. bryosalmonae cells were infrequently detected in the skeletal musculature of exposed experimental fish up to 72 h post-exposure. The route of migration through the fish to the kidney and spleen was not determined. All fish exposed to infected, disrupted bryozoans for 10, 30 and 90 min and maintained for up to 8 weeks developed clinical PKD.     

The occurrence of proliferative kidney disease (PKD) in cultured and wild fish: further investigations

Abstract. In an investigation of the occurrence of proliferative kidney disease (PKD) in freshwater fish other than rainbow trout, 18 species of wild fish and seven species of fish raised in cultivation wore sampled from waters where the disease occurred annually in rainbow trout, Oncorhynchus mykiss (Richardson). Results revealed that certain wild stocks of brown trout. Salmo trutta L., grayling, Thymallus thymallus L., and pike, Esox lucius L., were infected with PKD, as were cultivated Atlantic salmon, Salmo salar L., parr, brown trout and char, Salvelinus alpinus (L.). Microscopical examination revealed the presence of the PKX cell in these species and also intraluminal protozoa possibly related to the PKX cell, which were not found in the rainbow trout. Other species of freshwater fish had myxosporidan infections but, unlike PKD infection, there was little host/parasite tissue response. The PKX cell as a myxosporidan stage is discussed and the presence of the disease in wild fish is reported.     

Rodlet cells in Murray cod,Maccullochella peelii peelii (Mitchell), affected with chronic ulcerative dermatopathy

We have previously identified an unknown cell type in the gills of Murray cod affected with chronic ulcerative dermatopathy (CUD), a condition that causes severe erosion of epidermis surrounding cephalic and lateral line sensory canals. The condition arises in aquaculture facilities that utilize groundwater, with the cause of the condition suggested to be an unknown contaminant(s). Light and transmission electron microscopy were used to characterize and quantify the unknown cells in CUD‐affected Murray cod. The cells were identified as rodlet cells and were characterized by their oval or round shape, basally located nucleus, thick fibrillar capsule surrounding the cell, and multiple rodlet sacs containing a central electron‐dense core within the cell. Rodlet cells were present in the gills, kidney and intestine of non‐CUD‐affected and CUD‐affected Murray cod; however, differences in the numbers were observed between the groups of fish. A significantly greater number of rodlet cells were observed in the gills and collecting ducts of CUD‐affected fish. This is the first report of rodlet cells in Murray cod, and we suggest that the increased rodlet cell numbers in CUD‐affected Murray cod may be in response to unknown water contaminant(s) present in the groundwater that give rise to CUD.     

A dual infection by infectious cuticular epithelial necrosis virus and a Chlamydia-like organism in cultured Litopenaeus vannamei (Boone) in Ecuador

During 1996, microscopic examinations of post larvae and juveniles of moribund Litopenaeus vannamei showed multifocal necrosis in the cuticular epithelial tissues. In addition to these severe degenerative alterations in the epithelial cells typical of infectious cuticular epithelial necrosis virus (ICENV), columnar cells of the epithelium displayed small round intracytoplasmic inclusions in the necrotic tissue. Examination by electron microscopy of affected tissues demonstrated prokaryotic organisms in the cytoplasm of epithelial cells delineated by a distinct cytoplasmic vesicle; the prokaryotic organisms were morphologically similar to the genus Chlamydia. The necrotic tissue also showed the presence of particles of ICENV; the double infection by two different organisms in cuticular epithelial cells has not been reported previously. Two distinct stages in the intracellular development of a Chlamydia‐like organism were recognized: (1) pleomorphic elementary bodies (EBs) that were spherical to oval were often observed in the process of division or in forming a common chain of three cells, the cells were surrounded by a rigid cell envelope and the presence of a cap or plaque hexagonally arrayed; (2) the reticular bodies (RBs) were forms often in the process of division. These cells had an electron‐dense cytoplasm and contained a loose network of nuclear fibrils and a more fragile cell envelope. Regardless of the development stages of the Chlamydia‐like organism within the cytoplasmic vesicles, ICENV particles were observed, either dispersed or in clusters, surrounded or inside the vesicles. The potential adverse impact of this dual infection on shrimp culture should be considered, especially in high‐density operations.     

Proliferative kidney disease in Switzerland: current state of knowledge

This study presents an overview of the distribution of proliferative kidney disease (PKD) in wild and farmed fish in Switzerland based on two sources: (1) the results of routine diagnostic work at the National Fish Disease Laboratory, and (2) the results of a country‐wide survey for PKD. The first case of PKD in Switzerland was diagnosed in 1979 and since 1981, a few cases have been found every year. Affected species were rainbow trout, brown trout and grayling from rivers and fish farms. Most of the diseased fish were found in the lower altitude regions of the country (Swiss midlands). A seasonal distribution of the cases was evident: PKD‐positive fish occurred only from June to November withprominent peaks in August and September. Among infected fish all length classes were present, but the highest numbers of affected fish were found in the length classes <10 cm and 16–20 cm. The evaluation of the samples from a survey of the occurrence of PKD in 2000/2001 revealed a much broader geographical distribution of the infection than expected from routine diagnostic observations. Fish from 56 of 139 rivers analysed by histological examination were positive for parasites. Fish from 131 of these sites were examined macroscopically. Of these, trout from 45 sites showed gross alterations indicative for PKD. Most of the positive fish were found in the Swiss midlands north of the Alps. Fish from six of 65 fish farms investigated were found to be PKD‐positive by histopathological analysis, and in four of these cases fish could be identified as infected on the basis of gross examination.     

患“白体病”罗氏沼虾腹部肌肉病变的超微结构

取患“白体病”罗氏沼虾 (Macrobrachiumrosenbergii)腹部的病变组织及其鳃、心肌、肝胰腺、生殖腺等器官和组织 ,经光镜和电镜制样观察 ,未发现病原体 ,表明该病为非病原体引起的疾病。光镜下的病变组织结构与正常组织相比无明显异常。但在电镜下 ,病变主要表现为肌肉细胞的线粒体和肌质网的变性、坏死。线粒体肿胀、空泡化 ,肌质网增多且形成许多内陷小泡 ,表明细胞处于缺氧和钙代谢紊乱状态 ,这可能与虾生存的环境和养殖条件有关。根据超微病理与组织学观察结果 ,认为“罗氏沼虾白体病”更确切的命名应为“罗氏沼虾肌肉细胞线粒体坏死症”     

Proliferative kidney disease and renal myxosporidiosis in juvenile salmonids from rivers in England and Wales

The results from a survey for proliferative kidney disease (PKD) and renal myxosporidiosis in wild salmonids from rivers in England and Wales are presented. One hundred and eighty-five salmon, Salmo salar , 235 brown trout, Salmo trutta , 16 charr, Salvelinus alpinus, and five grayling, Thymallus thymallus, were obtained from 23 locations on 16 rivers between July and October 1997. They were examined for the presence of clinical PKD and for histological evidence of infections with Tetracapsula bryosalmonae and other renal myxozoans. Prevalence of infection with T. bryosalmonae detected histologically in brown trout varied from 11 to 43% in enzootic rivers and was only found in salmon in two rivers at low prevalence. Nephromegaly was positively associated with PKD in brown trout but in salmon mild nephromegaly was only associated with infection with an unidentified Chloromyxum sp.     

Ultrastructure of the macrogamont of Eimeria funduli, a coccidium parasitizing killifishes

Abstract. The ultrastructurc of the macrogamont of Eimeria funduli was studied in hepatocytcs of the killifish Funldulus grandis and F. similis. The macrogamont developed in a parasitophorous vacuole (PV) that was bounded by two unit membranes probably derived from an envelope of modified host rough cndoplasmic reticulum (RER). The membrane adjacent to the host-cell cytoplasm was lined with ribosomes. whereas the membrane facing the PV was smooth and blebbed into the PV. Intravacuolar tubules were not observed in the PV. The macrogamonts had a single limiting membrane with lew micropores. Developing macrogamonts had abundant RER. few Golgi bodies, and a few small polysaccharide granules. Nuclei of developing macrogamonts had dispersed chromatin and compact nucleoli. Polysaccharide granules in developing and mature macrogamonts were identified with the periodic acid-thiocarbohydrazide-silver proteinate technique. In mature macrogamonts, polysaccharide granules as large as 1 –2 μm in diameter nearly filled the cytoplasm. The cytoplasm contained three kinds of inclusions that were associated with either the RER or Golgi apparatus. Inclusions identified as type I wall-forming (WF) bodies were small, dense, membrane-bound granules that developed from the Golgi apparatus. Inclusions identified as type II WF bodies were usually dense, irregularly shaped bodies that occupied RER cisternae. Some type II WF bodies had a heterogeneous content but most were homogeneous. A third type of inclusion was related to the cytoplasmic aspect of RER cisternae and consisted of tubules and vacuoles enclosed in a membrane.     

“红肉病”文蛤中寄生类立克次体的超微结构与细胞病理学

报道了患“红肉病”文蛤的消化盲囊上皮中所发现的类立克次体以及所引起的受感染细胞的病理学变化。该病原包涵体呈球形或椭球形,有强嗜碱性和嗜酸性两种,电镜下观察到上皮细胞内充满了大量的类立克次体,其形态结构及大小表现了极大的多样性,受感染的细胞具有明显的病理学变化,内质网膨大为潴泡状,核糖体脱落;线粒体溶解或固缩,核膜肿胀,染色质固缩并出现液泡结构,高度感染时,细胞核与大部分的细胞器消失,在上皮细胞中,观察到类立克次体以二分裂和出芽方式进行繁殖。     

A brown trout (Salmo trutta) population faces devastating consequences due to proliferative kidney disease and temperature increase: A case study from Austria

Proliferative kidney disease (PKD) is an emerging parasitic disease that affects wild and farmed salmonid fishes worldwide. Currently, it is widespread in Europe with records in many countries. This study assessed the status of PKD in the river Wulka, a small Austrian river, which was known to host a vital brown trout population despite having high water temperatures. Contrary to the initial hypothesis that the river was free from PKD, we observed an overall PKD prevalence of 92%. Noninfected fish were restricted to the uppermost river section. Twenty-two out of 87 PKD-positive fish of all age classes showed clinical signs, while five individuals exhibited signs of recovering from PKD infection. The severity of clinical signs was significantly dependent on the fish length and age, with smaller or younger individuals being more severely affected. Comparison of data from the earlier fish sampling events with those results from 2017 and 2018, together with the analyses of the pattern of water temperature since 2010, indicates that Wulka's brown trout population might become extinct in the near future.     

皱纹盘鲍眼部组织的显微及亚显微结构观察

采用组织学和电镜的方法,对皱纹盘鲍(Haliotis discus hannai Ino)的眼部组织进行了光镜和电镜观察,以期为进一步从分子生物学角度解析鲍对光照的生理响应机制提供组织学与细胞学基础。结果发现,眼部组织由外至内依次为视网膜色素上皮细胞层、外核层、光感受器内节、内核层、黑色素颗粒沉积层、视觉纤维层。组织表面布满乳头状突起,每一乳状突起的顶端均具有一簇或两簇纤毛环。疏松结缔组织和平滑肌纤维等是组织内的主要成分,结缔组织间分布的胶原纤维等对保持细胞的弹性和韧性具有重要的作用。研究结果显示了鲍的眼部组织在感受和辨识外界光环境因子中的重要作用,也为鲍养殖生产中的光环境因子优化和调控、深入探讨鲍对光照的生理应答机制提供了形态学依据。