Maturation of circulating red blood cells in young baltic salmon (Salmo salar L.)

The maturation of circulating red blood cells (RBC) in salmon (Salmo salar L.) has been studied. A developmental RBC series of 6 classes based on morphological criteria was proposed. After a single dose of iron (55Fe) given to 5-6 months old salmon the incorporation of radioactivity in maturing RBC was followed by autoradiography on blood smears. The relative distribution of labelled RBC between the 6 classes in specimens taken 15, 20, 41, and 52 days after the injection of iron showed that the RBC matured along the proposed series. The projected area of the RBC increased by 100% during the maturation. This RBC growth was caused by an elongation of the RBC. Thus the ratio between short and long axes of the RBC decreased with maturation and can be used as a measure of the degree of maturation. The uptake of radioactive iron measured as grain count per RBC or per projected area of the RBC increased during the maturation in classes II-IV. Radioactive iron was not incorporated by mature RBC. The maturation time from class I to class VI was shorter than 41 days. RBC iron bound to non-haemoglobin proteins as a possible restriction in estimations of the haemoglobin concentrations of developing RBC is discussed.     

Histochemical localization of carbonic anhydrase in the pig's exocrine pancreas

Carbonic anhydrase is essential for the pancreatic secretion of NaHCO3. To localize the distribution of carbonic anhydrase in the exocrine gland and, hence, identify the potential cellular source of secreted NaHCO3, histochemical staining of carbonic anhydrase was carried out on pancreatic biopsies obtained from six pigs at both supramaximal intravenous secretin stimulation and at secretory rest. Tissue staining, using Hansson's technique, revealed a strong staining both in membranes and cytoplasm in the duct cells and a weaker cytoplasmic staining in the acinar cells. Staining reaction was abolished by 10(-5) mol l-1 acetazolamide. Duct cells, accordingly, seem to be responsible for NaHCO3 secretion, while acinar cells appear unlikely to contribute substantially to NaHCO3 secretion.     

Studies on the parasite fauna of juvenile Atlantic salmon,Salmo salar L., cultured in fresh water in eastern Scotland

The parasite fauna of juvenile Atlantic salmon,Salmo salar L., at a smolt-rearing unit in eastern Scotland was monitored from two weeks after first-feeding until smolting occurred (May 1973–April 1975). A total of 625 fish was examined. Wild salmon from the river supplying the unit with unfiltered water were also examined.The cultured fish yielded eight protozoan and seven metazoan parasite species, all derived from wild fish. The parasite fauna of cultured fish became established quickly, subsequent changes involving loss of some species and increasing abundance of others. The metazoan parasite fauna was restricted to those species with free-swimming infective stages capable of entering the tanks.     

Coronary artery lesions in Atlantic salmon (Salmo salar)

One hundred and fifteen hearts and dorsal aortas were studied in anesthetized or moribund salmon for presence of histological alterations associated with spawning and changing environmental conditions. The control group consisted of 26 fish hatched and raised under experimental and controlled conditions. The study group included 61 prespawners and 23 postspawners. The lesions observed were divided into three grades in increasing order of severity. They consisted of focal or diffuse intimal proliferation of cells resembling smooth muscle cells and frequent alteration of the underlying elastica. The changes were more frequently seen in larger branches of coronary arteries, arterial conus, and atrioventricular sulcus regions. Both the relative incidence and severity of lesions appeared greater in prespawners as compared to postspawners. This study indicates a high incidence of coronary lesions in spawning Atlantic salmon. In addition, the severity of the lesions appeared reduced in postspawners exposed to prolonged starvation. It is suggested that the Atlantc salmon with its exposure to varying environmental conditions may be a useful model to study the epidemiology of coronary artery disease.     

Histochemical localization of carbonic anhydrase in flask cells of the clawfrog mesonephros (Xenopus laevis Daudin) (author's transl)

Carbonic anhydrase activity was demonstrated in the flask cells of intermediate segments and collecting tubules of the clawfrog-mesonephros with the cobalt-bicarbonate method of H?usler (1958) and Hansson (1967). Additional the luminal surface of proximal and distal convoluted tubules reacted positively. By electron microscopy the enzyme activity in the flask cells was localized mainly on the surface of microvilli like cytoplasmic processes which lined the lumen of intracellular secretion capillaries. The existence of carbonic anhydrase in the flask cells is related to a possible physiological function of these cells.     

Genomic survey of early responses to viruses in Atlantic salmon, Salmo salar L

Viral diseases are one of the main problems and risk factors in aquaculture. At present diseases are diagnosed by detection of pathogens and clinical symptoms. Identification of genes involved in early responses to viruses is important for better knowledge of antiviral defence and development of diagnostic tools. The aim of this study was to search for gene markers common for viral infections in Atlantic salmon based on microarray analyses of a wide range of samples. Gene expression profiles from fish and cell cultures infected with different viruses and treated with the synthetic double-stranded RNA poly(I:C) were compared in order to identify virus responsive genes (VRG). The list of VRG defined in this study contained 117 genes with known or unidentified functions. Several genes, including the most highly ranked one (receptor transporting protein), had not been previously reported to be involved in antiviral defence. VRG were characterized by a rapid induction and low tissue specificity, and their expression levels were related to the viral load. Immunofluorescence analyses of proteins encoded by VRG in cardiac tissue of salmon with the viral disease cardiomyopathy syndrome (CMS) revealed a common expression pattern. In head kidney leukocytes VRG showed comparable or equal responses to CpG and poly(I:C), which mimic respectively bacterial DNA and viral RNA. Most VRG showed highly correlated expression with interferon-a (IFNa). Sequence comparison of salmon VRG with those from other species gave an understanding of the evolution of these genes, which showed a remarkably rapid sequence divergence in comparison with the entire proteome. VRG emerged both before and after separation of teleosts and tetrapods, and among genes found exclusively in fish species there were members of several multigene families: tripartite motif proteins, gig1- and gig2-like proteins. Several VRG, including genes with unknown functions and orthologs to mammalian RNA helicase RIG-I and chemokine C-X-C type 10, were present in cyprinid and salmonid fish but not in the phylogenetically advanced orders, suggesting that they have been lost in the evolution of Teleostei. Apparently, a number of genes involved in antiviral responses in salmon have acquired different functional roles in higher vertebrates.     

Cloning and expression analysis of an Atlantic salmon (Salmo salar L.) tapasin gene

Loading of the major histocompatibility complex (MHC) class I molecule with peptide is mediated by the multimeric peptide-loading complex in the ER where the glycoprotein tapasin (TAPBP) is required for stabilization of the complex and for control of peptide loading onto MHC class I. To expand our knowledge on antigen presentation genes in Atlantic salmon, we isolated a full-length salmon tapasin cDNA sequence (Sasa-TAPBP). It encoded a 443 bp amino acid sequence with two N-glycosylation sites, two conserved mammalian tapasin signature motifs, two Ig superfamily (IgSf) domains, a transmembrane (TM) domain and an ER-retention KK motif at the C-terminal end, indicative of a similar function as mammalian tapasins. We analysed the regulation of Sasa-TAPBP under immunostimulatory conditions and found an mRNA-upregulation during early infectious salmon anemia virus (ISAV) infection and poly I:C stimulation in vivo and in vitro, in line with our previous findings for other MHC class I pathway genes.     

Purification and partial characterization of an IgM-like serum immunoglobulin from Atlantic salmon (Salmo salar)

An IgM-like immunoglobulin was isolated from pooled sera collected from healthy Atlantic salmon. The immunoglobulin was purified by means of gel filtration followed by ion-exchange chromatography. It eluted from the ion-exchange column as two distinct peaks, but the two IgMs seems to be identical in their molecular natures except for net charge. The molecular weight of the unreduced (native) IgM was determined to be approximately 800 kilo Daltons (kD) when estimated by dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). However, when the molecular weight of the native IgM was estimated by gel filtration, a molecular weight of 1000 kD was obtained. Furthermore, the molecular weights of the heavy and the light chains were estimated by SDS-PAGE analysis to be about 72 and 27 kD respectively. The amount of IgM was found to range from 80 to 130 mg/100 ml serum. Isoelectric focusing demonstrated that the major part of the IgM molecules focused between pH 5 and pH 6.     

Blood volume changes and arginine vasotocin (AVT) blood concentration in conscious fresh water and salt water adapted ducks

Blood volume changes consisting in the removal and reinfusion respectively, of 10% of the estimated blood volume (23.2 ml on average) were induced to determine their effects on the blood concentration of arginine vasotocin (AVT), the antidiuretic hormone (ADH) of birds, in fresh water adapted ducks (water ducks) with blood osmolalities and ADH concentrations similar to those of normally hydrated mammals, and in salt water adapted ducks (salt ducks) with chronically elevated blood osmolalities and ADH concentrations. The investigations were carried out in steady state conditions, when infusion of 1 ml·min^–1 of isotonic saline was matched by the excretion in water ducks and when infusion of 0.4 ml·min^–1 of 1,000 mosmolal saline was matched by the salt gland excretion in the salt ducks.After blood removal, AVT blood concentration (mean ±SE) increased from 6.5±0.4 to 8.4±0.6 pg·ml^–1 in water ducks and from 18.1±1.6 to 22.6±1.9 pg·ml^–1 in salt ducks. The respective blood osmolalities of 297.4±1.4 and 318.6±3.3 mOsm·kg^–1 did not change. Reinfusion of the blood after steady-state conditions had been reattained decreased blood AVT from 7.9±0.7 to 6.7±0.5 pg·ml^–1 in water ducks. In the salt ducks AVT concentration had already returned to the control level before blood reinfusion which induced no further reduction. The blood osmolalities remained unchanged in both groups.During blood removal and reinfusion, the excretion rate of the kidneys in water ducks and the salt glands in salt ducks were temporarily reduced and enhanced respectively, the effect being not symmetrical for salt gland secretion.For water ducks, the volume sensitivity of AVT release was found comparable to that of mammals, when related to the induced blood volume changes, the responses to blood removal and reinfusion being approximately equal in absolute terms. In the salt ducks, the volume sensitivity of AVT release was clearly expressed during blood removal but insignificant during blood reinfusion.Supported by the Alexander v. Humboldt-StiftungSupported by the Deutsche Forschungsgemeinschaft     

Localization of natriuretic peptides in the cardiac pacemaker of Atlantic salmon (Salmo salar L.)

This study describes the location of the primary pacemaker at the sino-atrial (SA) junction and the localization of salmon cardiac peptide (sCP) and ventricular natriuretic peptide (VNP) in Atlantic salmon (Salmo salar L.). The pacemaker tissue appeared lightly stained and composed of: (1) wavy nerve bundles with oval elongated wavy appearing nuclei with pointed ends, (2) ganglion cells (12–22 μm) with granular cytoplasm and (3) wide muscle fibers with large nuclei (modified cardiomyocytes) clearly distinguishing them from the other myocardial cells. Pacemaker tissue was further evaluated using immunohistochemical staining. Immunoreactivity of natriuretic peptides (sCP and VNP) antisera showed specific staining in pacemaker ganglion cells in addition to the cardiomyocytes. Positive staining with anti-CD3? antisera in the pacemaker ganglion cells is a novel finding in teleosts and is consistent with observations in mammals. In conclusion, the Atlantic salmon pacemaker was shown to be located at the SA node and to harbor sCP and VNP peptides, suggesting a possible neuromodulatory and/or neurotransmitter role for these cardiac hormones within the teleost heart.     

Vibrio salmonicida pathogenesis analyzed by experimental challenge of Atlantic salmon (Salmo salar)

Cold-water vibriosis (CV) is a bacterial septicemia of farmed salmonid fish and cod caused by the Gram-negative bacterium Vibrio (Aliivibrio) salmonicida. To study the pathogenesis of this marine pathogen, Atlantic salmon was experimentally infected by immersion challenge with wild type V. salmonicida and the bacterial distribution in different organs was investigated at different time points. V. salmonicida was identified in the blood as early as 2 h after challenge demonstrating a rapid establishment of bacteremia without an initial period of colonization of the host. Two days after immersion challenge, only a few V. salmonicida were identified in the intestines, but the amount increased with time. In prolonged CV cases, V. salmonicida was the dominating bacterium of the gut microbiota causing a release of the pathogen to the water. We hypothesize that V. salmonicida uses the blood volume for proliferation during the infection of the fish and the salmonid intestine as a reservoir that favors survival and transmission. In addition, a motility-deficient V. salmonicida strain led us to investigate the impact of motility in the CV pathogenesis by comparing the virulence properties of the mutant with the wild type LFI1238 strain in both i.p. and immersion challenge experiments. V. salmonicida was shown to be highly dependent on motility to gain access to the fish host. After invasion, motility was no longer required for virulence, but the absence of normal flagellation delayed the disease development.     

CpG oligodeoxynucleotides and plasmid DNA stimulate Atlantic salmon (Salmo salar L.) leucocytes to produce supernatants with antiviral activity

Unmethylated CpG dinucleotides are more frequent in the genomes of bacteria and viruses than of vertebrates. We report herein that plasmid DNA and synthetic oliogodeoxynucleotides (ODNs) containing unmethylated CpG induce production of antiviral cytokine activity in Atlantic salmon leucocytes, whereas ODNs with an inverted motif (GpC) or with methylated cytosines have nearly no stimulatory effect. The adherent cell population, representing mainly macrophages, is directly activated by CpG-ODN, while the effect on the non-adherent population is weak. Since the peak antiviral activity in ODN-stimulated leucocytes is seen after 48h, this might indicate that the unmethylated DNA stimulates the adherent cells to produce co-stimulatory molecules, which in turn stimulates production of antiviral cytokines in the non-adherent cell population. The potent immune activation by CpG ODNs points to possible new applications as adjuvant in fish vaccines.