Utilization of molecular methods to relate Red Mark Syndrome affecting Oncorhynchus mykiss to an unculturable Rickettsiales organism

Red Mark Syndrome (RMS) is an inflammatory skin condition related to the presence of an unculturable Rickettsiales organism that affects farmed rainbow trout, Oncorhynchus mykiss. This disease is responsible of economic losses for fish farmers in Europe. The aim of this work was to confirm the presence of Rickettsiales organisms in affected fish and to develop a specific and sensitive system that can detect the unculturable Rickettsiales organisms in farmed trout. Trout gathered from affected and unaffected farms were initially submitted to necropsy and subsequently tissue samples were collected from each fish. Specific primers for a nested PCR (RiFCfw and RiFC), and a DNA probe for dot blot assay were designed using the 16S rRNA sequence. The use of the DNA probe to target amplicons as the template increased the sensitivity to 0.5 pg/\u3bcl DNA. Fifteen trout were analysed, of which nine showed visible skin lesions. Eight trout out of the nine with skin lesions were positive for the presence of the unculturable Rickettsiales organism DNA sequence. The tests developed can help rainbow trout producers to quickly treat fish in order to reduce economic losses. Up to 30% of fish with skin lesions are downgraded and rejected at the market by consumers