Soybean meal alters autochthonous microbial populations, microvilli morphology and compromises intestinal enterocyte integrity of rainbow trout, Oncorhynchus mykiss (Walbaum)

Rainbow trout were fed either a diet containing fishmeal (FM) as the crude protein source or a diet containing 50% replacement with soybean meal (SBM) for 16 weeks. An enteritis-like effect was observed in the SBM group; villi, enterocytes and microvilli were noticeably damaged compared with the FM group. The posterior intestine microvilli of SBM-fed fish were significantly shorter and the anterior intestine microvilli significantly less dense than the FM-fed fish. Electron microscopy confirmed the presence of autochthonous bacterial populations associated with microvilli of both fish groups. Reduced density of microvilli consequently led to increased exposure of enterocyte tight junctions, which combined with necrotic enterocytes is likely to diminish the protective barrier of the intestinal epithelium. No significant differences in total viable counts of culturable microbial populations were found between the groups in any of the intestinal regions. A total of 1500 isolates were tentatively placed into groups or genera, according to standard methods. Subsequent partial 16S rRNA sequencing revealed species that have not been identified from the rainbow trout intestine previously. Compared with the FM group levels of Psychrobacter spp. and yeast were considerably higher in the SBM group; a reduction of Aeromonas spp. was also observed.     

Development and characterization of a cell line TTCF from endangered mahseer Tor tor (Ham.)

Tor tor is an important game and food fish of India with a distribution throughout Asia from the trans-Himalayan region to the Mekong River basin to Malaysia, Pakistan, Bangladesh and Indonesia. A new cell line named TTCF was developed from the caudal fin of T. tor for the first time. The cell line was optimally maintained at 28°C in Leibovitz-15 (L-15) medium supplemented with 20% fetal bovine serum (FBS). The propagation of TTCF cells showed a high plating efficiency of 63.00%. The cytogenetic analysis revealed a diploid count of 100 chromosomes at passage 15, 30, 45 and 60 passages. The viability of the TTCF cell line was found to be 72% after 6 months of cryopreservation in liquid nitrogen (-196°C). The origin of the cell lines was confirmed by the amplification of 578- and 655-bp sequences of 16S rRNA and cytochrome oxidase subunit I (COI) genes of mitochondrial DNA (mtDNA) respectively. TTCF cells were successfully transfected with green fluorescent protein (GFP) reporter plasmids. Further, immunocytochemistry studies confirm its fibroblastic morphology of cells. Genotoxicity assessment of H?O? in TTCF cell line revealed the utility of TTCF cell line as in vitro model for aquatic toxicological studies.     

Fish-based methods for assessing European running waters: a synthesis

Abstract  The European Union, Water Framework Directive (WFD) requires monitoring of riverine fish fauna. When the WFD came into force in 2000, most of the EU member states did not have fish-based assessment methods compliant to WFD requirements. Therefore, the objectives of FAME ( http://fame.boku.ac.at ), a project under the fifth R&D Framework Programme of the European Commission were to develop, evaluate and implement a standardised fish-based method for assessing the ecological status of European running waters. This paper synthesises the outputs of FAME and defines future research needs. Two different methodologies were used: the so-called spatially based modelling and the site-specific modelling, the latter leading to the European Fish Index (EFI). The advantage of the EFI is that, despite being a single index, it is applicable to a wide range of environmental conditions across Europe precluding the need for inter-calibration. The EFI will support the WFD towards harmonised/standardised assessment and management of running waters in Europe, thus enabling comparative analyses of the ecological status of running waters across Europe.     

Immunohistochemical study on changes in gill Na+/K+-ATPase α-subunit during smoltification in the wild masu salmon, Oncorhynchus masou

Changes in immunoreactivity of Na⁺/K⁺-ATPase -subunit in gill sections of wild masu salmon (Oncorhynchus masou) during the parr-smolt transformation (smoltification) were compared with changes in gill Na⁺/K⁺-ATPase specific activity. Gill Na⁺/K⁺-ATPase specific activity increased from April and peaked in May. Immunohistochemical analysis, using an antiserum against a synthetic oligopeptide based on the conserved region of the Na⁺/K⁺-ATPase -subunit, revealed that immunoreactivity was confined to chloride cells in the surface layer of primary lamellae and the proximal end of secondary lamellae. The size and number of these cells increased gradually from February to May; however, the number of chloride cells of the secondary lamellae decreased in May. These data suggest that the synthesis of Na⁺/K⁺-ATPase and the proliferation of chloride cells occur prior to the elevation of enzyme activity. Moreover, it is likely the proliferation and hypertrophy of chloride cells on primary lamellae prepare smolts for entry into seawater and migration in the ocean.     

Insulin and IGF-I receptors and tyrosine kinase activity in carp ovaries: changes with reproductive cycle

Insulin and insulin-like growth factor I (IGF-I) receptors from carp ovaries were semipurified with wheat germ agglutinin at different moments of the reproductive cycle and their binding characteristics and tyrosine kinase activity were studied. Specific receptors for insulin and IGF-I were found. IGF-I receptors presented higher binding (23.8 ± 1.5%), number of receptors (965 ± 20fm/mg) and affinity (K_D 0.24 ± 0.03nM) than those shown for insulin receptors (4.1 ± 1%, 530 ± 85fm/mg and 0.85 ± 0.1nM, respectively). Insulin and IGF-I receptors have a tyrosine kinase activity which is not different from that found in muscle of the same species. Seasonal changes were found in binding, with maximum values for insulin and IGF-I reached at the end of pre-spawning period (June). However, while IGF-I binding was observed in all stages, insulin binding decreased in autumn and disappeared in winter, which suggests a different role for the two peptides in ovarian physiology.     

Evaluation of Streptocephalus dichotomus nauplii as a larval diet for freshwater prawn Macrobrachium rosenbergii

This study was performed to assess the nutritional value of Streptocephalus dichotomus nauplii and compared with standard larval diet and parthenogenetic strain of Artemia nauplii. The effectiveness of live feed was determined by feeding freshwater prawn Macrobrachium rosenbergii postlarvae. Results on the growth rate, weight gain, survival, fatty acid and amino acid composition show a significant variation between the dietary treatments. Mean larval growth was significantly different ( P  < 0.001) between control diet (13.5 ± 0.5 mm) and live feeds ( Streptocephalus nauplii; 18.4 ± 0.5 mm and Artemia nauplii 18.7 ± 0.2 mm). The weight of larvae-fed Streptocephalus (41.1 ± 1.44 mg) and Artemia nauplii (41.7 ± 0.2 mg) was not significantly different; however, treatment with live feeds was significantly higher than the control (16.3 ± 0.5 mg) ( P  < 0.001). Proximate composition on the fairy shrimp reveals that they are rich in protein, lipid, essential amino acids (EAA) and essential fatty acids (EFA). The polyenoic unsaturated fatty acid (18 : 2 n-6 and 18 : 3 n-3) and highly unsaturated fatty acid (20 : 4 n-3 and 20 : 5 n-3) show a dramatic increase in larval tissue relative to its proportional composition in the live diets. Amino acid composition in the live feeds, Streptocephalus and Artemia nauplii, perfectly reflects on the amino acid proportions in the larvae-fed diets which confirm its utilization.     

Design and validation of a RT‐qPCR procedure for diagnosis and quantification of most types of infectious pancreatic necrosis virus using a single pair of degenerated primers

Infectious pancreatic necrosis virus (IPNV) is an important virus which affects the salmonid aquaculture industry worldwide; therefore, it is important to develop rapid and reliable methods of diagnosis to detect the disease at early stages. Nowadays, RT‐qPCR is replacing other methods because it provides additional information on the viral load, which is important to have a better understanding of the virus replication level and of the stage of the infection and its risk level. The main problem stems from the high diversity of this virus, which can compromise the reliability of the diagnosis. In this study, we have designed an RT‐qPCR procedure for diagnosis and quantification of IPNV based on a single pair of primers targeted to segment B. The procedure has been validated, in vitro and in vivo, testing two different types of standards against seven reference strains and 23 field isolates from different types. The procedure is reliable for the detection of any type, with a detection limit of 31 TCID₅₀ mL^?1, 50 pfu mL^?1 or 66 RNA copies mL^?1, depending on the standard. All the standard curves showed high reliability (R² > 0.95). The results support the high reliability of this new procedure for the diagnosis and quantification of IPNV.     

Inoculation of ‘Golden Delicious’ Apple Trees on M9 Rootstock with Azotobacter Improves Nutrient Uptake and Growth Indices

ABSTRACT

Roots of young ‘Golden Delicious’ apple on M9 rootstock were inoculated with four strains of Azotobacter chroococcum, which were isolated from various soils. Effects of these strains in combination with different levels of nitrogen (N) fertilizer and compost on plant growth and nutrient uptake were studied over two seasons. Therefore, a factorial arrangement included four strains of A. chroococcum, two levels of N-fertilizer (0 and 35 mg N kg^?1soil of ammonium nitrate) and two levels of compost (0 and 12 g kg^?1 soil of air-dried vermicompost). Among the four strains, AFA₁₄₆ was the most beneficial strain, as it increased leaf area, leaf potassium (K), magnesium (Mg), iron (Fe), manganese (Mn), zinc (Zn), and boron (B) uptake and root N, phosphorus (P), potassium (K), Mn, and Zn. The combination of AFA₁₄₆ strain, compost and N fertilizer increased leaf uptake of Ca, Mg, Fe, Mn, Zn, and B, and root uptake of P, K, Ca, Mg, Mn, and copper (Cu), and root dry weight.