Application of molecular techniques to identify immune/stress associated gene transcripts in bluefin tuna and their subsequent in vitro expression

Bluefin tuna (Thunnus spp.) are a globally threatened genus of fish that currently constitutes one of the most economically important food fisheries in the world. Intensive culture efforts have intensified in recent years to maximize fishery profitability and mitigate market dependency on wild stock; yet little is currently known concerning health and immune functions of these highly specialized organisms and best culture practices continue to be investigated. In an attempt to aid future aquaculture interests and to provide general tools for identifying immune responses and disease status for this group of fishes, we have investigates a number of immune and diseaseidentifying aspects regarding bluefin tuna through the use of molecular laboratory techniques centering on quantitative PCR. In this presentation, we will focus on our identification of 20 immune, stress, or growth related mRNA sequences of bluefin and subsequent in vitro explorations regarding the utility of selected immune and stress associated genes as biomarkers in bluefin through gene expression analysis. Our findings suggest that temperature exerts influence in the timing but not the degree of an innate inflammatory response in bluefin tuna and that different cell populations have differential responsiveness to heat shock in this heterothermic species. Further, E. coli LPS stimulation of Southern bluefin leukocytes failed to induce host Hsp70; yet, increased inflammatory signaling was observed in following a combined heat shock/LPS stimulation, suggesting a time specific ‘cross-talk’ between these signaling pathways during immune and stress associated responses. A strong correlation between Hsp70 and IL-8 transcriptional expression was also observed following LPS/heat shock stimulation of leukocytes and five potential heat shock response elements were subsequently identified on the gene promoter region of IL-8 indicating that heat shock co-activation of this chemokine previously identified in mammals is also likely present in fish.