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Arctic charr ( Salvelinus alpinus) are the northernmost distributed freshwater fish and can grow at water temperatures as low as 0.2 °C. Other teleost species have impaired immune function at temperatures that Arctic charr thrive in, and thus, charr may maintain immune function at these temperatures. In this study, a fibroblastic cell line, named ACBA, derived from the bulbus arteriosus ( BA) of Arctic charr was developed for use in immune studies at various temperatures. ACBA has undergone more than forty passages at 18 °C over 3 years, while showing no signs of senescence-associated β-galactosidase activity and producing nitric oxide. Remarkably, ACBA cells survived and maintained some mitotic activity even at 1 °C for over 3 months. At these low temperatures, ACBA also continued to produce MH class I proteins. After challenge with poly I:C, only antiviral Mx proteins were induced while MH proteins remained constant. When exposed to live viruses, ACBA was shown to permit viral infection and replication of IPNV, VHSV IVa and CSV at 14 °C. Yet at the preferred temperature of 4 °C, only VHSV IVa was shown to replicate within ACBA. This study provides evidence that Arctic charr cells can maintain immune function while also resisting infection with intracellular pathogens at low temperatures. [ABSTRACT FROM AUTHOR]

A cell line, WE-cfin11e, with an epithelial-like morphology was developed from a caudal fin of walleye, Sander vitreus ( Mitchill), characterized as distinct from the established walleye caudal fin fibroblast-like cell line, WE-cfin11f, and compared with WE-cfin11f for susceptibility to VHSV IVb. Immunocytochemistry and confocal microscopy were used to localize the intermediate filament protein, vimentin, the tight junction protein, zonula occludens-1 ( ZO-1), the extracellular matrix protein, collagen I, and the viral protein, G. Although both cell lines contained vimentin, only WE-cfin11e stained for ZO-1 and only WE-cfin11f stained for collagen I. Ascorbic acid increased the accumulation of collagen I and caused the appearance of collagen fibres only in WE-cfin11f cultures. At 14 °C, both cell lines produced VHSV IVb, but the infection developed more rapidly in WE-cfin11f. At 4 °C, both cell lines became infected with VHSV IVb as judged by the expression of viral proteins, N and G, but only WE-cfin11f produced virus. The results suggest that cold temperatures can modulate viral tropism. [ABSTRACT FROM AUTHOR]

Compared to fathead minnow, walleye demonstrate low susceptibility to experimental infection with VHSV IVb, regardless of route of exposure or water temperature at time of infection. In triplicate and duplicate groups, walleye were intraperitoneally (i.p.) injected (102-108 pfu/fish) or waterborne-exposed (w; 1.4 × 107 pfu mL−1) with VHSV IVb. High cumulative mortality (64-100%) and severe gross lesions associated with VHSV IVb infection were evident only in fish i.p. injected with 108 pfu at 12 °C. These fish had multifocal necrosis of several tissues including the gill and heart. There was no difference in mortality between walleye infected (w or i.p.) at 12 °C (spring stocking) compared with a declining temperature profile from 18 to 12 °C (fall stocking). There were significant differences ( P < 0.05) in mortality between four extant walleye strains following i.p. infection, indicating that the choice of walleye strain for stocking might be an important consideration. Viral antigen was found in both i.p. and w-exposed walleye using immunohistochemistry, mostly within the gill and skin of w-exposed fish and most prominently in dermal fibrocytes. VHSV IVb was detected in multiple tissues from 6 to 21 days post-infection using reverse transcriptase quantitative polymerase chain reaction (RT- qPCR). [ABSTRACT FROM AUTHOR]

A cell line, WE-cfin11f, with a fibroblast-like morphology was developed from a walleye caudal fin and used to study the intersection of thermobiology of walleye, Sander vitreus ( Mitchill), with the thermal requirements for replication of viral haemorrhagic septicaemia virus ( VHSV) IVb. WE-cfin11f proliferated from 10 to 32 °C and endured as a monolayer for at least a week at 1-34 °C. WE-cfin11f adopted an epithelial shape and did not proliferate at 4 °C. Adding VHSV IVb to cultures at 4 and 14 °C but not 26 °C led to cytopathic effects ( CPE) and virus production. At 4 °C, virus production developed more slowly, but Western blotting showed more N protein accumulation. Infecting monolayer cultures at 4 °C for 7 days and then shifting them to 26 °C resulted in the monolayers being broken in small areas by CPE, but with time at 26 °C, the monolayers were restored. These results suggest that at 26 °C, the VHSV IVb life cycle stages responsible for CPE can be completed, but the production of virus and the initiation of infections cannot be accomplished. [ABSTRACT FROM AUTHOR]

A letter to the editor related to fomites for viral hemorrhagic septicemia virus, is presented.

A novel fibroblast-like cell line RTHDF was established from hypodermal connective tissue of rainbow trout Oncorhynchus mykiss and telomerase activity was demonstrated early and late in cell line development. When RTHDF cells were exposed to bioenergetic stress, i.e. anoxia, activation of the stress activated member of the mitogen-activated protein kinase family, p38MAPK and induction of heat shock protein (Hsp70) were evident. The time-course of the p38MAPK activation and the induction of Hsp70 expression in RTHDF were studied in response to chemically induced anoxia. p38MAPK was activated rapidly, with maximal activity after 10 min of anoxia. Hsp70 was induced after 30 min of anoxia, followed by overnight recovery in growth medium at 21° C. Using the p38MAPK-specific inhibitor SB203580, the enhanced expression of Hsp70 occurred independently of p38MAPK activation in RTHDF. These data suggests that RTHDF can be useful in studying biochemical responses of teleost cells to environmental stress. [ABSTRACT FROM AUTHOR]

Cortisol at 1000 and 100 ng/ml, and less consistently at 10 ng/ml, inhibited increases in cell number and 3H-thymidine incorporation by cultures of the rainbow trout (Oncorhynchus mykiss) monocyte/macrophage cell line, RTS11. Cell viability was not altered by cortisol, although a small decline in the capacity of cultures to reduce the redox dye, Alamar Blue was observed. In cortisol-treated cultures, more round and fewer spread cells were evident. Similar results were observed with dexamethasone but not cortisone. The glucocorticoid receptor antagonist, RU-486, prevented the effects of cortisol on RTS11 proliferation, and shape. In co-culture with the spleen stroma cell line (RTS34st) or in medium conditioned by RTS34st, the proliferation of RTS11 was enhanced. Treating RTS11/RTS34st co-cultures or RTS11 cultures in RTS34st conditioned medium with cortisol did not inhibit RTS11 proliferation. Overall these experiments suggest that proliferation of rainbow trout macrophages is regulated by cortisol, but the effect is modulated by the cellular micro-environment, possibly through the release of cytokines. [Copyright &y& Elsevier]

An outdoor microcosm study was conducted in order to evaluate the kinetics of immunomodulation in rainbow trout (Oncorhynchus mykiss) exposed to liquid creosote. Fish were sampled on d 7, 14, 21, and 28 from microcosms dosed initially with 0, 3, and 10 µl/L of creosote. Pronephros leukocytes were monitored for phagocytic activity, oxidative burst, and surface immunoglobulin-positive (SIg[sup +]) B-cell counts. Oxidative burst was inhibited by creosote exposure; however, by sampling d 28, the burst response returned to near control levels. Phagocytic activity, on the other hand, was stimulated, peaking on sampling d 7, then returning to near control levels by d 28. Although control SIg[sup +] B-cell counts were quite variable across sampling days, SIg[sup +] B-cell counts were also elevated in creosote-exposed fish after seven days of exposure. SIg[sup +] B-cell numbers decreased significantly to near control levels during the remainder of the study. The overall results from this study confirm that creosote has the potential to alter certain immune parameters, and emphasize the importance of monitoring changes in the immune system during exposure. Polycyclic aromatic hydrocarbons (PAHs), a major constituent of liquid creosote, are the suspected immune-altering agents. [ABSTRACT FROM AUTHOR]

Screening for environmental contaminants in milk is generally conducted by chemical analysis, yet such an approach may be time-consuming and expensive, and is not indicative of the physiological consequences of such exposure. The focus of this review is to summarize those constituents of milk that may be altered by maternal exposure to one of the most biologically active environmental pollutants, 2,3,7,8-tetrachlorodibenzo-p-dioxin, and to develop the case for their use as biomarkers of response. Several chemical and/or cellular components of milk are potentially useful as biomarkers, and may be developed as convenient, biologically relevant indicators of maternal exposure to dioxinlike compounds. [ABSTRACT FROM AUTHOR]

Collagenase was used to prepare primary cell cultures from rainbow trout, Oncorhynchus mykiss (Walbaum), gills. Although difficult to subcultivate, one primary culture led to the development of a cell, RTgil1-W1. RTgiIl-W1 grew in the basal medium, L-15, supplemented with foetal bovine serum at between 5 and 10%, but not in L-15 alone. The cells have been passaged approximately 50 times over a 4-year period. At confluency, the cell shape was predominantly polygonal or epithelial-like. RTgill-W1 cultures were demonstrated by DNA staining with H33258 and by growth on agar to be contaminated with mycoplasma, but this contaminant was eradicated by treatment with mycoplasma removal agent (MRA) and BM cyclin. [ABSTRACT FROM AUTHOR]