Immunity augmentation in Botia dario (Hamilton, 1822) through carotenoid: a dietary approach.

The immunity and health status of ornamental fish is an important aspect, as they are kept in a confined environment and various stressful conditions which lead to depletion of overall colourful appearance and mortality. The carotenoids can act as immunity boosters in captive aquarium system and may be supplemented in the feed as aquarium fish have no access to natural carotenoids. The study aimed to assess the role of carotenoid on the immunity of B. dario. Marigold petal meal is an important source of carotenoids and used in experimental diets. Four immunogenes namely IL20, TLR9, TRAIL, and Nramp in B. dario were characterized and also studied for their relative expression in the kidney after feeding the fish with marigold petal meal supplemented diet. The expression pattern of the genes was compared with the fish of nature. The IL20 and Nramp gene were upregulated significantly (p < 0.05) in the fish of nature as compared to the experimental fish at the 60th day of feeding carotenoid-rich diet. But the TLR9 and TRAIL gene was upregulated significantly (p < 0.05) in experimental fish as compared to nature. The haematological parameters of fish after feeding with the experimental enriched diet for 60 days also confirmed the role of carotenoids in immunity.

Effect of feeding Lipopolysaccharide as an immunostimulant on immune response and immune gene expression of Labeo bata.

This study investigates the effects of dietary lipopolysaccharide (LPS) as an immunostimulant on hematology, innate immunity, immune gene expression and protection against Edwardsiella tarda on Labeo bata. A basal diet supplemented with 0, 50, 100 and 150mg LPS kg-1diet was fed to the four different groups for 30days. The haematological (total erythrocyte count, total leukocyte count, total serum protein, albumin and globulin), innate immune parameters (respiratory burst, serum lysozyme, myeloperoxidase and serum bactericidal activity), immune gene expression (C3, ß-2 microglobulin, lysozyme g, transferrin, IFN-1, IFN-γ) were monitored at 7th, 15th, 30th day and one day post challenge (DPC) with E. tarda. All the studied haematological, innate immune parameters and expression of immune gene increased significantly (p≤0.05) in LPS fed group in comparison with control. However the group fed 100mgkg-1 LPS in feed showed highest activity on 7th day and 1DPC. The group fed 100mgkg-1 LPS also recorded highest relative percent survivability after challenge with E. tarda. Therefore this study suggests that LPS at 100mgkg-1 could be used as an immunostimulant in feed to enhance the protection of bata during periods of increased disease risk.

Innate immune response of Indian major carp, Labeo rohita infected with oomycete pathogen Aphanomyces invadans.

The fish pathogenic oomycete Aphanomyces invadans is the causative agent of epizootic ulcerative syndrome (EUS), a fish disease of international significance and reportable to the World Organisation for Animal Health. In spite of the current and potential impact of A. invadans infection on fisheries and aquaculture sectors of the world, very little is known about the host-A. invadans interactions. In the present study, following experimental infection with A. invadans in one of the Indian major carps, Labeo rohita, sequential changes in various innate immune parameters were monitored. The results indicated that at early stages of infection, no significant changes in any of the studied innate immune parameters were observed. However, at the advanced stages of infection from 6 to 12 days post infection (dpi), the respiratory burst and alternate complement activity were significantly higher whereas lysozyme, antiproteases and α-2 macroglobulin values were significantly lower than the control group and also from the infected group at earlier stages of infection. Since, the possibility of vaccination of fish against A. invadans appears remote due to difficulties in eliciting a specific antibody response, the information generated in the present study could be useful for developing strategies for improving resistance to A. invadans infection by stimulating the innate immunity through immunomodulation.

Molecular characterization and expression of type-I interferon gene in Labeo rohita.

Genes coding for type-I interferon (I-IFN) has been cloned from Labeo rohita, a commercially important and widely cultured fish in India and South East Asia. In the present study, full-length gene of I-IFN was amplified and sequenced. The sequence analysis revealed that I-IFN consists of 1,786 bp genomic sequence with four introns and five exons and an ORF of 546 bp encoding for a putative protein of 181 amino acids. The mature protein has a molecular weight of 18.97 kDa and consists of 158 amino acids and a signal peptide of 23 amino acids at the N terminus. The sequence carries I-IFN signature motif, one glycosylation site, two conserved cystine amino acids and other conserved amino acids. The sequence showed highest similarity to that of Cyprinus carpio (84%). In silico analysis of the rohu I-IFN protein was done using various bioinformatic tools. The constitutive expression of I-IFN gene was found to be more in spleen compared to gill and kidney in real time PCR assay. Expression of I-IFN increased about 20-fold in cultured kidney cell 2 h after induction with poly I:C and showed maximum expression at 8 h post-induction.

DNA fingerprinting of Flavobacterium columnare using RAPD-PCR.

In the present study, DNA fingerprinting of eight strains of Flavobacterium columnare was done by random amplification of polymorphic DNA (RAPD) fingerprinting method. The strains were collected from Fish Health Management Division, Central Institute of Freshwater Aquaculture, Bhubaneswar, India. A total number of 160 primers were screened for RAPD-PCR, of which 10 primers yielded amplification with all the strains. The molecular weight of amplified bands varied from 0.29-2.63 Kb. The number of bands varied from 1 to 8. Unique band was seen with primer OPY-15 with molecular weight 0.75 Kb that can be used for epidemiological study. Genetic variability was investigated using NTSYS software. Highest genetic similarity was found between MS1 and MS3 followed by MS5 and MS7. Minimum genetic similarity was found between MS2 and MS8. Phylogenetic tree was constructed using UPGMA and neighbor joining methods.