'No-mucus skin disease' of rainbow trout, Oncorhynchus mykiss (Walbaum): a case report

Abstract A skin disease of intensively reared salmonids in Ontario hatcheries, known to the farmers as'no-mucus skin disease', is reported for the first time. It was characterized by erosive and ulcerative lesions found mainly on the flanks of fingerlings, which resulted in exposure of the tips of the scales. Associated with these lesions were colonies of bacteria seen in the SEM to be clustered round the mucous cell pores and under-running the margins of epithelial cells. The cause of this condition is unknown, although the response of fish to formalin treatment, and the presence of bacilli seen in skin scrapings and in the SEM, suggest that bacteria are responsible.     

Aeromonas salmonicida: determination of an antigen associated with protective immunity and evaluation of an experimental bacterin

Abstract. Protection against Aeromonas salmonicida was determined by passive immunization and with various bacterin preparations. Rabbit antiserum was prepared against a rough, virulent strain of A. salmonicida (AS-1R), the same strain boiled (AS-1R, boiled), and an avirulent, smooth strain of this same isolate (AS-1S). Cross-absorption, cross-passive protection and analysis by counter immunoelectrophoresis of various extraction methods were studied. It was shown that AS-1R cells contained an additional antigen not present in AS-1R (boiled) and AS-1S cells. Antiserum to the AS-1R antigen passively protected sockeye salmon, Oncorhynchus nerka (Walbaum), against a virulent challenge, and antisera to AS-1R (boiled) and AS-1S were not protective. The antigen was not destroyed by formalin or heat at 5°C for 60 min, but it appeared to be partially inactivated with proteolytic enzymes. The antigen was produced in casein yeast beef (CYB) broth up to 32 h but not thereafter, and low yields were obtained in tryptic soy or brain heart infusion (BHI) broth. It was extracted from cells with ethylenediamine tetraacetic acid (EDTA) and especially alkaline hydrolysis, but not with proteolytic enzymes or detergents. The detergents appeared to destroy the antigen. We concluded that the antigen was protein and is most likely the external A-protein (AP) reported for rough, virulent strains of A, salmonicida. Various methods of preparing A. salmonicida bacterins were evaluated by determining the level of protective immunity induced in intraperitoneally (i.p.) vaccinated fish. Growth of cells in CYB or BHI broth resulted in production of only rough (autoagglutinated in saline) variants of A. salmonicida. Although only rough variants were associated with protective immunity, one strain was not protective, it was avirulent by bath challenge. Bacterins prepared in CYB were more efficacious than those grown in BHI, but inactivation with formalin, iodine, or glutaraldehyde worked equally well. However, boiling the bacterin or filtering the cells from the bacterin removed its efficacy. Methods of releasing the AP were evaluated by sonification, pH-lysis, disaggregation and treatment with EDTA, and all treatments worked equally well. Also, precipitation on to aluminium or use of Freund's complete adjuvant did not significantly improve the protection. In parenterally vaccinated fish, protection was demonstrated by challenging the fish at various levels by bath, injection or cohabitation with infected fish. The best protection was demonstrated using the cohabitation challenge method. The potency and field efficacy of an A. salmonicida bacterin prepared in CYB broth and extracted with 5 mM EDTA was evaluated. Fish were vaccinated by i.p, injection and potency was determined in the laboratory by experimental challenge and in the field by natural challenge. Chinook salmon, O.ishawytschu (Walbaum), developed immunity within seven days at 10°C. The bacterin could be diluted up to 1:2000 without loss of potency. The field tests results were equivocal; however, (he prevalence of infection was lower in vaccinated fish.     

Ecological and landscape effects on genetic distance in an assemblage of headwater fishes

Environmental divergence along hierarchically structured longitudinal gradients may constitute barriers to gene flow in river networks for headwater specialised species. While known, this phenomenon has not been well studied, especially with regard to degree of headwater specialisation. We examined six headwater species that differ in habitat specialisation to assess whether patterns of differentiation vary according to geographic or environmental distance. We also identified regional environmental or anthropogenically induced fragmentation effects by comparing within‐drainage patterns of genetic distance across replicate watersheds. We used a comparative modelling framework to determine whether isolation by distance or isolation by resistance of large river habitats was a better predictor of genetic distance across species. The influence of reservoir presence and regional network characteristics that may influence the hydrology and size of large river habitats were also assessed. Resistance effects from large rivers were closely related to headwater specialisation, with increased specialisation leading to increased resistance and loss of drainagewide population connectivity. These results affirm that dendritic networks naturally fragment headwater specialised species. Further isolation from anthropogenic fragmentation was detected in two of the six drainages, indicating interactions with system‐specific conditions. Landscape variables related to the hydrology of large rivers also affected genetic distance in predicted ways, supporting the importance of large rivers in genetically structuring headwater species in drainage networks.     

A rapid immunoperoxidase-based virus neutralization assay for salmonid alphavirus used for a serological survey in Northern Ireland

A modified virus neutralization (VN) assay was developed to replace an existing assay read on the presence or absence of virus-induced cytopathic effect (CPE). The modified assay used a monoclonal antibody to salmon pancreas disease virus as the first layer of an immunoperoxidase (IPX)-based immunostaining technique to detect viral growth. The IPX-based VN assay required only 3 days to perform, and the adoption of a 96-well microtitre format facilitated a high throughput of samples requiring small volumes of serum, cells and virus. When 352 sera from farmed salmon and 302 sera from farmed trout were tested by both the modified and the original CPE-based assays, overall correlations of 97.72 and 96.03% were, respectively, obtained (96.94% combined). When the modified assay was used to test 188 sera collected from wild salmonids in freshwater river systems in Northern Ireland, no positive results were recorded.     

Use of collagen shields for treatment of chronic bilateral corneal ulcers in a pet rabbit

A four-year-old, 1·3-kg entire male Holland Lop pet rabbit (Oryctolagus cuniculus) was presented with a two-day history of bilateral epiphora and blepharospasm. Fluorescein staining revealed bilateral medio-ventral corneal ulceration. Initial topical treatment included ophthalmic broad-spectrum antibiotics and artificial tear solutions. Over a six-month period, antibiotics were changed based on corneal culture and sensitivity test results, and periodic corneal debridement was performed. With little to no improvement, and recurrence of the previous clinical signs, surgical intervention was considered but withheld because of individual pet consideration and owner's reluctance. Hydrophilic contact lenses were placed for corneal support but failed and caused a severe ocular response. Corneal collagen shields were placed bilaterally in order to promote corneal healing. Recheck examination performed seven days after placement revealed complete resolution of the ulcer in one eye and major reduction of the corneal ulcer in the contra-lateral eye.     

IHHN virus: Infectivity and pathogenicity studies in Penaeus stylirostris and Penaeus vannamei

Infectivity studies of infectious hypodermal and hematopoietic necrosis virus (IHHNV) were conducted on Penaeus stylirostris and P. vannamei (representing species highly susceptible and highly refractory to the disease) via intramuscular injections of the virus. Distinctive histological lesion patterns were observed between species. Six target organ systems were compared interspecifically. The gills and nerve cord/ganglia were shown to undergo significantly higher degrees of tissue damage in infected P. stylirostris compared with infected P. vannamei. Pathogenetic differences correlated well with observed differences in epizootiology between the two species.The infectivity study confirmed that IHHN disease is virus-caused. Cell-free extracts from IHHNV-infected shrimp, when injected into healthy, susceptible shrimp, produced mortalities and Cowdry type A intranuclear eosinophilic inclusion bodies presently considered pathognomonic for the disease.     

Evaluation of interferon gamma (IFN-γ) of <Emphasis Type="Italic">Labeo rohita</Emphasis> as an immunomodulator: in vitro expression model

Interferon gamma (IFN-γ) or type II interferon is a cytokine that is critical for innate and adaptive immunity against viral and some bacterial and protozoal infections. The importance of IFN-γ in the immune system lies in its ability to inhibit viral replication directly and most importantly from its immunomodulatory effects. Previously, we successfully co-administered IFN-γ along with GAPDH gene of Edwardsiella tarda as bicistronic DNA vaccine in Labeo rohita. In order to ascertain the individual role of IFN-γ, the present study involves cloning and expression of 552-bp IFN-γ open-reading frame (ORF) of L. rohita in striped snakehead (SSN-1) cell line using eukaryotic expression vector system (pQE-TriSystem) followed by transfection in peripheral blood lymphocytes (PBMCs) to evaluate its immunomodulatory ability in comparison to polyinosinic-polycytidylic acid (Poly I:C)-treated PBMCs. The 18.7-kDa protein, expressed in the pQE-IFNγ-transfected SSN-1 cells, reacted with anti-His antibody in Western blot confirming it to be recombinant IFN-γ, whereas the relative expression of IFN-γ, iNOS, Mx, and IL-1β genes in PBMCs was quantified at 24 h and 48 h post treatment by qPCR. The comparative kinetics of all four genes showed significantly (p?<?0.05) high upregulation pattern in both pQE-IFNγ-transfected cell group and Poly I:C-treated cell group demonstrating recombinant IFN-γ as an equally efficient inducer like Poly I:C. Thus, our in vitro experiment results highlight the immunomodulatory potential of recombinant IFN-γ as an analogue to synthetic Poly I:C which warranted future studies to further explore the potential of recombinant IFN-γ as an effective vaccine adjuvant against different microbial invasion.     

Growth, digestive enzyme activities and hepatic glycogen levels in juvenile Colossoma macropomum Cuvier from South America during feeding, starvation and refeeding

Abstract. Feeding trials with juvenile Colossoma macropomum Cuvier (approximately 40g) were carried out to study their protein requirements and their efficiency in metabolizing vegetable protein. Two groups of diets were used. In one group, the crude protein consisted exclusively of fish protein concentrate. In the second group the protein consisted of a mixture (55:45) of vegetable and animal protein without fish meal. In each group 30 and 50% protein levels were used. The highest weight increase (1·64%/day) and the lowest feeding rate were obtained with the 50% protein fish meal free diet. During the trial all fish accumulated visceral fat while body protein content decreased and dry matter increased. There was a high correlation between liver weight and liver glycogen. The highest amounts of liver glycogen were induced by diets containing fish meal and/or low protein content. In contrast lo visceral fat deposits, accumulated liver glycogen was of less significance for surviving starvation periods. The activity of digestive enzymes depends on both stomach and gut fullness. Trypsin activity, contrary to pepsin, was correlated with diet protein content, a-amylase activity was not correlated with diet composition.     

Mating and spawning of freshwater crayfish, Austropotamobius pallipes Lereboullet, under laboratory conditions

Abstract. The study includes mating and spawning data of freshwater crayfish, Austropotamobius pallipes Lereboullet, obtained in four consecutive years (1987–1990). A total number of 586 wild-caught female A. pallipes in the 21·65 mm to 45·60mm carapace length (CL) size were held under laboratory conditions at different dates in September–October. Initial densities at mating time were 21 and 24 crayfish/m² with a 1:2 male:female ratio. High percentages of mating were obtained (97·4–100%) except in 1989 (57·5%). In all years, most of the matings (90%) were concentrated in a period of 8–12 days with mean water temperatures between 12°C and 13·5°C. Spawning took place a few days after mating (minimum 2 days, maximum 14 days) at mean water temperatures between 8·2°C and 10·8°C. In the first 3 years, almost 100% of mated females spawned. However, in the fourth year (1990), when the mean size of crayfish was smaller, only 72·4% of mated females spawned. The mean number of pleopodal eggs was 64 (maximum 220, minimum 18). Egg diameter ranged between 2·30 and 3·25 mm (mean value 2·78 mm). Pleopodal egg number was positively correlated with carapace length ( r = 0·72).