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Quantitation of immune response gene expression and cellular localisation of interleukin-1β mRNA in Atlantic salmon, Salmo salar L., affected by amoebic gill disease (AGD)
- Bridle, A.R., Morrison, R.N., Cunningham, P.M.C., Nowak, B.F.
- Veterinary immunology and immunopathology 2006 v.114 no.1-2 pp. 121-134
- Salmo salar, salmon, immune response, gene expression, quantitative analysis, interleukin-1, messenger RNA, protein synthesis, amebiasis, Amoebida, fish diseases, reverse transcriptase polymerase chain reaction, nucleic acid hybridization, genes, nitric oxide synthase, animal proteins, gills, liver, kidneys, gene expression regulation, infection
- The characterisation of selected immune response genes during amoebic gill disease (AGD) in Atlantic salmon, Salmo salar L., was performed using semi-quantitative RT-PCR, quantitative real-time RT-PCR (qRT-PCR), and in situ hybridisation (ISH). The immune response genes of interest were interleukin-1β (IL-1β), inducible nitric oxide synthase (iNOS), serum amyloid A (SAA), and serum amyloid P-like pentraxin (SAP). Atlantic salmon were inoculated with the ectoparasite Neoparamoeba sp., the causative agent of AGD, and gill, liver and anterior kidney tissue sampled at 0, 7 and 14 d post-inoculation (p.i.). Semi-quantitative RT-PCR was performed on the tissue samples to identify up/down-regulated mRNA expression relative to uninfected control fish and normalised to the housekeeping gene, β-actin. Interleukin-1β (IL-1β) was the only immune response gene of those investigated whose mRNA was differentially regulated in any of the tissues and was found to be up-regulated in the gills by semi-quantitative RT-PCR. Increased gill IL-1β mRNA expression was then accurately quantitated and confirmed using probe-based qRT-PCR. The cellular localisation of the IL-1β mRNA expression in the gills of uninfected and infected fish was then determined by ISH using an IL-1β-specific biotinylated cRNA probe. Expression of IL-1β mRNA was localised to filament and lamellar epithelium pavement cells in gills of uninfected and infected Atlantic salmon. These data implicate the involvement of IL-1β at the site of infection, the gills, of Atlantic salmon during AGD. This work supports previous studies that suggest IL-1β is important in the regulation of the fish immune response to parasitic infection but additionally shows the cellular localisation of fish IL-1β mRNA expression during infection.