Tenacibaculum maritimum, a member of the phylum Bacteroidetes, is responsible for the farmed-fish disease tenacibaculosis, which is generating significant economic losses to the rapidly growing aquaculture industry. Diseased fish characteristically exhibit external symptoms such as skin ulcers, mouth erosion, fin necrosis and rotted tails.
Despite, the economic importance of T. maritimum, there is limited knowledge about its pathogenicity mechanisms and relevant virulence factors. Gram-negative pathogens secrete virulence factors that play pivotal roles in pathogenesis, including extracellular lytic enzymes and toxins such collagenases, hemolysins and proteases. Collagenases are important virulence factors that disrupt extracellular host structures during infection, a virulence mechanism applied by several pathogenic Bacteroidetes such as the better-studied cold-water disease agent Flavobacterium psychrophilum.
The genomes of multiple T. maritimum strains sequenced in our group show that they harbor several genes encoding secreted virulence factors, including collagenases and a homologue gene to a TcB-TcC component of known tripartite toxin complex (Tc toxin). The collagenases identified from T. maritimum are multi-domain proteins and possess the collagenase catalytic motif HExxH. The gene encoding for the TcB-TcC homologue gene is located adjacent to multiple hypothetical cluster of genes located in the same operon[SM1] . We studied the protein profile of the secretome obtained from T. maritimum and were able to show that the secretome is indeed packed with potential virulence factors including extracellular lytic enzymes such as collagenases, a potential Tc-toxin subunit a TcB-TcC component and multiple proteins encoded by the cluster of genes belonging to the operon harboring the TcB-TcC component. In addition, a prelimenary collagenase assay performed on gelatin from fish skin showed the presence of enzymes with collagenase activity in the protein cocktail of the secretome.
The aim of this project is to characterize and determine the roles of potential virulence factors such as collagenases and a homologue of TcB-TcC component of a Tc toxin complex (and optionally additional secreted enzymes) in the pathogenesis of T. maritimum using molecular and/or [SM2] biochemical methods. For this purpose, we will recombinantly produce the protein candidates and perform in vitro assays to study their mechanisms, which will give insights into the potential use of these proteins for the development of vaccines and drugs against tenacibaculosis.
Methods to use
Gene cloning
Protein expression, extraction and purification
Biochemistry/Enzymology
Potentially (depending on the interests of the candidate): Crystallography or in vivo infection assays
Supervisors: Prof. Dirk Linke, Dr. Sophanit Mekasha